Systems for producing multilayered particles, fibers and sprays and methods for administering the same

ABSTRACT

Capsules and particles with at least one encapsulated and/or entrapped agent, such as therapeutic agent, imaging agents, and other constituents may be produced by electrohydrodynamic processes. More particularly, the agent encapsulated in a vehicle, capsule, particle, vector, or carrier may maximize treatment and/or imaging of malignant cancers while minimizing the adverse effects of treatment and/or imaging.

CROSS-REFERENCE TO RELATED APPLICATIONS

This application claims priority to and the benefit of U.S. ProvisionalApplication Ser. No. 60/746,311, filed May 3, 2006, and U.S. ProvisionalApplication Ser. No. 60/886,225, filed Jan. 23, 2007, the disclosures ofwhich are expressly incorporated herein by reference in their entirety.

BACKGROUND

1. Field of the Invention

The invention relates to systems and methods for producing capsules andparticles with at least one encapsulated and/or entrapped agent, such astherapeutic agent, imaging agents, and other constituents. Moreparticularly, the agent encapsulated in a vehicle, capsule, particle,vector, or carrier may maximize treatment and/or imaging of malignancywhile minimizing the adverse effects of treatment and/or imaging.

2. Related Art

Cancer is a class of diseases or disorders characterized by uncontrolleddivision of cells and the ability of these to spread, either by directgrowth into adjacent tissue through invasion, or by implantation intodistant sites by metastasis. Cancer may affect people at all ages, butrisk tends to increase with age. It is one of the principal causes ofdeath in developed countries.

For example, according to the National Cancer Institute, breast canceraffected between 12 and 13 per every 10,000 women in 2003. Althoughimaging and early diagnostic tools have been improving over the past twodecades, current breast cancer early detection is far from infalible,especially when using mammograms in younger women. While magneticresonance imaging (MRI and ultrasound) and laser-based imagingtechniques have been used or evaluated, the issue for best-possiblecontrast between healthy and cancerous tissue for any imaginingtechnique is a longstanding one.

A current challenge facing scientists is determining how to design atherapeutic or an imaging agent and its vehicle, vector or carrier inorder to maximize treatment and imaging of malignant cancers in patientswhile minimizing the adverse effects of treatment. Moreover, theselective delivery of therapeutic agents to a desired part of the bodyis also a nontrivial issue. Current treatments may lead to insufficienttumor distribution or therapeutic agents and often cause adverse effectson patients. Systemic injections of therapeutic agents carryconsequences associated with their nonspecific dispersion in the bodyand have a limited therapeutic agent distribution throughout thetargeted malignancy. One approach to overcome these short comings is todesign an effective therapeutic or imaging agent delivery vehicle bymaking vehicles or capsules containing the desired therapeutic agent.

Formation of vehicles or capsules that are small enough to be deliveredinto the human body by means of inhalation, injection or permeationthrough the skin has received significant attention. The outer skin orshell of the vesicles may be chemically functionalized with receptorsand other species to selectively target certain organs. There areseveral methodologies available, such as electrospray and two-capillaryjet systems to fabricate small vesicles.

Emulsion-polymerization technologies, such as DC coaxial electrospray,AC coaxial electrospray, and electrohydrodynamics (EHD) are well knownmethods that may produce micron-sized capsules. In general, a solutioncontaining the target compound or compounds to be encapsulated, alsocalled “encapsulates” is emulsified into another fluid to solutionhaving at least one substance capable of forming a shell or envelopearound the encapsulate dispersed droplets. Althoughemulsion-polymerization methods are relatively scalable, there areseveral limitations associated with the methods, such as the inabilityto encapsulate the target in a quantitative manner, and the high-shearproduction of emulsions may compromise the integrity of mechanicallydelicate encapsulates, such as biological constituents such as proteins,genetic material, and other molecules of biological origin.

Coaxial electrospray based on application of DC electrical potentialsbetween a coaxial capillary fixture delivering theencapsulate-containing core liquid and a shell-forming liquid precursor,and a collector surface or counter-electrode, has the ability to producevesicles in the micrometer and sub-micrometer range. Although DC coaxialelectrospray may be relatively gentle to biological encapsulates, itsmajor limitation that it lacks simplicity in equipment, design,scalability, and thus cost effectiveness.

As an alternative to DC based electrospray, AC electrospray may beemployed. For example, AC electrospray may be used to produceencapsulates entrapped within a bioabsorbable biopolymeric matrix, suchas polylactic acid. AC electrospray yields essentially an electricallyneutral electrospray. While there are advantages associated with chargeneutrality, such as decreased ability of the particles or electrospraydroplets to absorb indiscriminately over non-targeted surfaces, andavoidance of a potential charge buildup problem. One disadvantage of ACelectrospray is that it produces undesirablely large particles havingsizes well above one micron. For many medical applications, such aspenetration of the blood brain barrier, AC electrospray derivedparticles are unacceptably large.

In addition to electrospray methodologies, coaxial liquid jet systemcombined with sol-gel chemistry, such as EHD, may be employed tofabricate vesicles or capsules. Use of coaxial, two-capillary coaxialarrangements to simultaneously deliver two fluids in the presence ofelectric field gradients are well known in the art.

Briefly, in this method, the chemistry and physical properties of thetwo fluids and the values at variables such as electric field strengthand flow rates of the two fluids may determine the structure of mattercollected onto a collector electrode, which may be located at a distancefrom the exit region of the two-capillary coaxial arrangement. At theexit region, the compound two-fluid structure may form an electrifiedmeniscus that may adopt various shapes, such as s Taylor cone.

FIG. 1, which illustrates a fluid flow generated by a two-coaxialcapillary system of the prior art, depicts a two-fluid stream 100 in thepresence of electric field gradients, where the internal fluid 104 isenveloped by the external fluid 106. The quasi-conical Taylor conestructure 108 issues an electrified compound two-fluid jet from its apex110. Here, the electrified liquid jet experiences thinning due tosame-charge repulsion effects. Moreover, the thinning of the jet may bea function of the physical properties of the two liquids such as,dielectric constants, viscosities, conductivities and surface tensions.Although differentiated two-fluid structures may occur when fluid 104and fluid 106 do not mix, they may also occur when fluid 104 and fluid106 are miscible or partially miscible, because both fluids flow underthe so-called laminar flow regime.

Laminar flows may be non-turbulent, which may minimize mixing betweenflowing fluid layers. Thus, since the two fluids may not mix to thepoint of forming a single fluid phase, the thinning electrifiedtwo-fluid jet may enter into a chaotic path resembling whippingphenomena. At a point along its path toward a collection zone orcollector body 112, the compound two-fluid jet may experience anelectrical charge oscillatory phenomenon known as Rayleigh instability.This may cause the compound two-fluid jet to no longer experienceprogressive thinning, but an oscillatory thinning and thickening regimewhich may eventually lead to jet breakup into a droplet-in-dropletregime, or compound electrospray regime.

The chemical and physical properties of the two fluids may be controlledto produce a variety of structures collected at the collection zone orcollector electrode 112. For example, if fluid 106 yields a solidstructure through solvent evaporation and precipitation of a solidphase, fluid 104 may be encapsulated into structures such as hollowfibers, hollow beaded fibers or capsules, for example. Alternatively,the chemical and physical properties of the two liquids may be adjustedto cause no solidification of fluid 104 and fluid 106, solidification ofone of the two fluids, or solidification of both fluids during the timeof travel of the compound charged structures from the two-fluidelectrified meniscus to the collection zone 112.

Referring to FIG. 1, regions 3, 4, and 5 are shown. If certainphysicochemical phenomena lead to solidification of fluid 106 in region3, tubular structures encapsulating fluid 104 inside a solid shellformed from fluid 106 may be obtained. If however, solidificationphenomena in fluid 106 occur in region 4, hollow beaded fibers withencapsulated fluid 104 may be obtained. Alternatively, if solidificationphenomena in fluid 106 occur in region 5, capsules with encapsulatedfluid 104 may be obtained. Wetted fibers, wetted beaded fibers or wettedparticles may result in regions 3, 4, and 5, respectively, in caseswhere fluid 104 solidifies but fluid 106 does not. Core-shell solidstructures may result when both fluids solidify prior to reaching thecollection zone.

Although coaxial two-capillary systems may be employed to produce thecore-shell structures, described above, there are several disadvantagesassociated with the conventional systems. In particular, when a direct,parallel scale-up of the process to increase process throughput isattempted a micro-fabrication problem occurs. For instance, a typicalrange of internal diameters for the inner and outer capillaries areabout 0.1 to about 0.3 mm and about 0.3 to about 1.0 mm, respectively.In order to build an instrument consisting of many such coaxial,two-capillary fixtures for scaled up production of a desired core-shellstructure, it is necessary to produce each individual fixture with innerand outer capillaries aligned as close to coaxial as possible, and alsowith high reproducibility in their diameters. With modernmicro-fabrication techniques such challenge may be met, however, thesetechniques are very complex and not cost effective.

In particular, a conventional way to manage fluid flow through manyorifices, capillaries, conduits or two-capillary coaxial fixtures is byusing one means for forcing flow through all same fluid orifices,capillaries, conduits or two-capillary coaxial fixtures, not bycontrolling the fluid flow rate through each individual fluid flow path.This is the reason why, for example, fabricating a parallel scaled upproduction of a desired core-shell structure is difficult and expensive.If there is variability in diameter from inner or outer capillary of onetwo-capillary coaxial fixture to another in excess of about 2% or 3%, itis not possible to produce a desired core-shell structure without theoccurrence of undesirable structures. With such prior art two-capillaryfixture, small differences in the overall pressure drop profiles of theinner and outer capillaries also causes undesirable effects.

SUMMARY OF THE INVENTION

The invention satisfies the above needs by providing systems and methodsfor fabricating capsules with entrapped or encapsulated therapeuticagents and/or imaging agents. More particularly, the systems and methodsof the invention permits a wider range of capsule size, keeps chemicalinteractions leading to degradation of therapeutic agents and/or imagingagents to sufficiently low and acceptable levels, involves lessmanufacturing steps, and is more cost effective than conventionalmethods.

According to one aspect of the invention, an electrohydrodynamic systemfor producing a capsule having at least one encapsulated agent isprovided. The system may include a hollow tube having an interiorconfigured to receive a core fluid, a fluid source surrounding saidhollow tube, a core fluid supply tube arranged to supply the core fluidto said interior of said hollow tube, a shell fluid supply tube arrangedto supply the shell fluid to said shell fluid source, and an electricpotential source to subject the core fluid and the shell fluid to anelectric potential to cause the fluids to form a jet including an atleast two-fluid electrically charged fluid. The system may include acore fluid reservoir. The system may also include a shell fluidreservoir. The encapsulated agent may be at least one of a therapeuticagent and an imaging agent.

In a particular aspect, the system may also include a collectorelectrode positioned above the fluid bath. Additionally, the system mayinclude an extractor body positioned between the fluid bath and thecollector electrode.

In a more particular aspect, the fluid source may be a fluid bath. Thefluid source may also be a porous material. The porous material may be asponge. Moreover, the fluid source may be a plurality of tubes.

According to one aspect of the invention, a system for producing acapsule having at least one encapsulated agent is provided. The systemmay include plurality of hollow tubes, an encasement surrounding saidplurality of hollow tubes, a core fluid supply tube arranged to supplythe core fluid to the interior of said plurality of hollow tubes, ashell fluid supply tube arranged to supply the shell fluid, and anelectric potential source to subject the core fluid and the shell fluidto a electric potential to cause the fluids to form a jet having an atleast two-fluid electrically charged fluid. The system may also includea collector electrode. Moreover, the system may also include anextractor body.

In a further aspect, the system of the invention may include a corefluid reservoir. Moreover, the system may also include a shell fluidreservoir.

In a specific aspect, the plurality of hollow tubes may becircumferentially arranged in said encasement. The plurality of hollowtubes may also be linearly arranged linearly arranged in said encasementcomprising at least two plates. The encasement may be configured toreceive and delivery the shell fluid. The shell supply tube may beconfigured and arranged to supply the shell fluid to a space between tosaid plurality of hollow tubes.

In a particular aspect, the system of the invention may be configured toperform upward flow electrohydrodynamics. Alternatively, the system ofthe invention may be configured to perform downward flowelectrohydrodynamics.

Additional features, advantages, and embodiments of the invention may beset forth or apparent from consideration of the following detaileddescription, drawings, and claims. Moreover, it is to be understood thatboth the foregoing summary of the invention and the following detaileddescription are exemplary and intended to provide further explanationwithout limiting the scope of the invention as claimed.

BRIEF DESCRIPTION OF THE DRAWINGS

The accompanying drawings, which are included to provide a furtherunderstanding of the invention, are incorporated in and constitute apart of this specification; illustrate embodiments of the invention andtogether with the detailed description serve to explain the principlesof the invention. No attempt is made to show structural details of theinvention in more detail than may be necessary for a fundamentalunderstanding of the invention and various ways in which it may bepracticed.

FIG. 1 is a schematic showing a two-fluid electrified jet, which is astrewn of fluids that may include at least two distinct fluid layers,according to principles of the invention.

FIG. 2 is a schematic showing the general architecture of a capsuleproduced by the methods of the invention.

FIG. 3 is a schematic showing an upward flow electrohydrodynamic system,according to principles of the invention.

FIG. 4 is a blow-up schematic of the upward flow electrohydrodynamicsystem of FIG. 3, showing the addition of the extractor.

FIG. 5 is a schematic showing a downward flow electrohydrodynamic systemusing a single capillary tube to form a two-fluid electrified jet,according to principles of the invention.

FIG. 6A is a schematic depicting a downward flow electrohydrodynamicsystem using a plurality of tubes or conduits to deliver the shellfluid, according to principles of the invention.

FIG. 6B is a cross-section of FIG. 6A.

FIG. 7A is a schematic showing a downward flow electrohydrodynamicsystem using a plurality of capillary tubes positioned between twoplates to deliver the core fluid, according to principles of theinvention.

FIG. 7B is a cross-section of FIG. 7A.

FIG. 8A is a schematic showing a downward flow electrohydrodynamicsystem using a plurality of capillary tubes to deliver the core fluid,according to principles of the invention.

FIG. 8B is a cross-section of FIG. 8A.

FIG. 9 is a schematic showing a downward flow electrohydrodynamic systemused to produce a capsule having at least three layers, according toprinciples of the invention.

FIG. 10 is a schematic showing a downward flow electrohydrodynamicsystem using a porous body to deliver the shell fluid to produce thetwo-fluid Taylor cone; according to principles of the invention.

FIG. 11 is a schematic of an upward flow electrohydrodynamic systemwhere the core fluid reservoir may rotate, according to principles ofthe invention.

FIG. 12 is a photograph of the electrifies meniscus formed under anelectric field and forced flow of the shell formulation.

FIG. 13A is a schematic depicting a method of the invention based on acombination of chemical reactions between the electrospray and itssurrounding gas phase, according to principles of the invention.

FIG. 13B is a schematic depicting a method of the invention based on acombination of chemical reactions between the electrospray and itssurrounding gas phase, according to principles of the invention.

FIG. 14 shows a photograph obtained by confocal microscopy. Panel I isthe unfocused signal and Panel II shows a focuses signal in the corefluid region of the capsule.

FIG. 15 is a graph showing the activity of transaminase in both itsencapsulated and in free solution, once rotation is converted to anequivalent D- (or L-) glutamine concentration via calibration curves.

DETAILED DESCRIPTION OF THE INVENTION

It is understood that the invention is not limited to the particularmethodology, protocols, and reagents, etc., described herein, as thesemay vary as the skilled artisan will recognize. It is also to beunderstood that the terminology used herein is used for the purpose ofdescribing particular embodiments only, and is not intended to limit thescope of the invention. It also is be noted that as used herein and inthe appended claims, the singular forms “a,” “an,” and “the” include theplural reference unless the context clearly dictates otherwise. This,for example, a reference to “a capsule” is a reference to one or morecapsules and equivalents thereof known to those skilled in the art.

Unless defined otherwise, all technical and scientific terms used hereinhave the same meanings as commonly understood by one of ordinary skillin the art to which the invention pertains. The embodiments of theinvention and the various features and advantageous details thereof areexplained more fully with reference to the non-limiting embodimentsand/or illustrated in the accompanying drawings and detailed in thefollowing description. It should be noted that the features illustratedin the drawings are not necessarily drawn to scale, and features of oneembodiment may be employed with other embodiments as the skilled artisanwould recognize, even if not explicitly stated herein.

Any numerical values recited herein include all values from the lowervalue to the upper value in increments of one unit provided that thereis a separation of at least two units between any lower value and anyhigher value. As an example, if it is stated that the concentration of acomponent or value of a process variable such as, for example, size,temperature, pressure, time and the like, is, for example, from 1 to 90,specifically from 20 to 80, more specifically from 30 to 70, it isintended that values such as 15 to 85, 22 to 68, 43 to 51, 30 to 32etc., are expressly enumerated in this specification. For values whichare less than one, one unit is considered to be 0.0001, 0.001,0.01 or0.1 a appropriate. These are only examples of what is specificallyintended and all possible combinations of numerical values between thelowest value and the highest value enumerated are to be considered to beexpressly stated in this application in a similar manner.

Moreover, provided immediately below is a “Definition” section wherecertain terms related to the invention are defined specifically.Particular methods, devices, and materials are described, although anymethods and materials similar or equivalent to those described hereincan be used in the practice or testing of the invention. All referencesreferred to herein are incorporated by reference herein in theirentirety.

Definitions

BSA is bovine serum albumin

CED is convection enhanced delivery

CFM is confocal fluorescence microscopy

EHD is electrohydrodynamics

PBS is phosphate-buffer saline

PEG is Poly(ethylene glycol)

PLA is poly(lactic acid)

PLC is polycaprolactone

PLGA is poly(lactic-co-glycolic acid)

The terms “active agent,” “drug,” “therapeutic agent,” and“pharmacologically active agent” are used interchangeably herein torefer to a chemical material or compound which, when administered to anorganism (human or animal) induces a desired pharmacologic effect.Included are derivatives and analogs of those compounds or classes ofcompounds specifically mentioned that also induce the desiredpharmacologic effect. In particular, the therapeutic agent may encompassa single biological or abiological chemical compound, or to acombination of biological and abiological compounds that may be requiredto cause a desirable therapeutic effect.

By “pharmaceutically acceptable carrier” is meant a material ormaterials that are suitable for drug administration and not biologicallyor otherwise undesireble, i.e., that may be administered to anindividual along with an active agent without causing any undesirablebiological effects or interacting in a deleterious manner with any ofthe other components of the pharmaceutical formulation in which it iscontained.

Similarly, a “pharmacologically acceptable” salt, ester or otherderivative of an active agent as provided herein is a salt, ester orother derivative that is not biologically or otherwise undesirable.

By the terms “effective amount” or “therapeutically effective amount” ofan agent as provided herein are meant a nontoxic but sufficient amountof the agent to provide the desired therapeutic effect. The exact amountrequired will vary from subject to subject, depending on the age,weight, and general condition of the subject, the severity of thecondition being treated, the judgment of the clinician, and the like.Thus, it is not possible to specify an exact “effective amount.”However, an appropriate “effective” amount in any individual case may bedetermined by one of ordinary skill in the art using only routineexperimentation.

The terms “treating” and “treatment” as used herein refer to reductionin severity and/or frequency of symptoms, elimination of symptoms and/orunderlying cause, prevention of the occurrence of symptoms and/or theirunderlying cause, and improvement or remediation of damage. Thus, forexample, the present method of “treating” individuals with cancer, asthe term “treating” is used herein, encompasses treatment of cancer in aclinically symptomatic individual.

The terms “condition,” “disease” and “disorder” are used interchangeablyherein as referring to a physiological state that can be detected,prevented or treated by administration of a therapeutic agent asdescribed herein. Exemplary diseases and conditions may include, but notlimited to, cancer such as breast cancer, glioma, pancreatic cancer,leukemia, and lymphoma.

The term “patient” as in treatment of “a patient” refers to a mammalianindividual afflicted with or prone to a condition, disease or disorderas specified herein, and includes both humans and animals.

The term “nucleic acid,” as used herein may include an oligonucleotide,nucleotide, or polynucleotide, and fragments thereof, and to DNA or RNAof genomic or synthetic origin which may be single- or double-stranded,and represent the sense or antisense strand, to peptide nucleic acid(PNA), to small interfering RNA (siRNA) molecules, or to any DNA-like orRNA like material, natural or synthetic in origin.

The term “transfection,” as used herein includes the process ofintroducing a DNA expression vector into a cell. Various methods oftransfection are possible including microinjection or lipofection.

The term “transformation” as used herein generally refers to a processby which exogenous DNA enters and changes a recipient cell. It may occurunder natural or artificial conditions using various methods well knownin the art. Transformation may rely on any known method for theinsertion of foreign nucleic acid sequences into a prokaryotic oreukaryotic host cell. The method is selected based on the type of heatcell being transformed and may include, but is not limited to, viralinfection, electroporation, heat shock, and lipofection.

Antisense gene: an antisense gene is constructed by reversing theorientation of the gene with respect to its promoter so that theantisense strand is transcribed.

Antisense RNA: an RNA molecule complementary to a particular RNAtranscript that can hybridize to the transcript and block its function.

The term “functional equivalent,” as used herein generally refers to aprotein or nucleic acid molecule that possesses functional or structuralcharacteristics that is substantially similar to a heterologous protein,polypeptide, enzyme, or nucleic acid. A functional equivalent of aprotein may contain modifications depending on the necessity of suchmodifications for the performance of a specific function. The term“functional equivalent” is intended to include the “fragments,”“mutants,” “hybrids,” “variants,” “analogs,” or “chemical derivatives”of a molecule.

“Compound electrified fluid jet” as used herein generally refers to astream of fluid composed of at least two distinct fluid layers. Forexample, when two fluids are used to form a compound electrified jet byEHD, the outer fluid is the fluid precursor for the shell of the capsulemade by the compound electrified fluid jet by EHD method and the innerfluid is the precursor for the core of the capsule.

According to the invention, an agent is “encapsulated” when the capsulehas at least one core region and at least one particle shell region in alayered architecture. The capsule may further contain at least oneagent, such as a therapeutic agent or an imaging agent.

According to the invention, an agent is “entrapped” when the agent isdispersed within a biocompatible matrix composed of one or morebiocompatible polymers, for example, and distinct onion-like layers arenot apparent.

The term “carcinoma” as used herein generally refers to malignant tumorsderived from epithelial cells. This group may represent the most commoncancers, including the common forms of breast, prostate, lung and coloncancer.

The terms “lymphoma” and “leukemia” as used herein generally refer tomalignant tumors derived from blood and bone marrow cells.

The term “sarcoma” as used herein generally refers to malignant tumorsderived from connective tissue, or mesenchymal cells.

The term “mesothelioma” as used herein generally refers to tumorsderived from the mesothelial cells lining the peritoneum and the pleura.

The term “glioma” as used herein generally refers to tumors derived fromglia, the most common type of brain cell.

The term “germinoma” as used herein generally refers to tumors derivedfrom germ cells, generally found in the testicle and ovary.

The term “choriocarcinoma” as used herein generally refers to malignanttumors derived from the placenta.

The term “attached,” as used herein generally refers to covalentbinding, adsorption, and physical immobilization. The terms “associatedwith,” “binding” and “bound” are identical in meaning to the term“attached.”

The term “nanoparticle” as used herein generally refers to a particle,generally metallic, semiconduction, magnetic, ceramic and dielectric,having a diameter in the range of about 1 nm to about 1000 nm.

The terms “polymer” or “biopolymer”, as used herein generally refer to acompound having two or more monomer units, and is intended to includelinear and branched polymers, and copolymers, the term “branchedpolymers” encompassing simple branched structures as well ashyperbranched and dendritic polymers. The term “monomer” is used hereinto refer to compounds that are not polymeric. “Polymers” or“biopolymers” herein may be naturally occurring, chemically modified, orchemically synthesized.

The term “functional group” generally refers to compounds that may besuitable for chemically binding of a first and second molecule together.Chemical bonding may be considered to broadly cover bonding with somecovalent character with or without polar bonding and can have propertiesof ligand-metal bonding along with various degrees of ionic bonding. Thefunctional group may refer to ligand-receptor binding relationships werecovalent bonding may not be the typical association. The functionalgroup may be selected based on the composition of the molecule. Anotherfunctional group of the linker may be suitable for covalent bonding afirst and second molecule together. Covalent bonding refers broadly tocovalent bonds with sigma bonds, pi bonds, hydrogen bonds, otherdelocalized covalent bonds and/or other covalent bonding types, and maybe polarized bonds with or without ionic bonding components and thelike. Covalent linkers include functionalized organic molecules. Thefunctional groups may include hydroxyl groups, amino groups, carboxylgroups, carboxylic acid anhydride groups, mercapto groups, andhydrosilicon groups.

The invention relates to systems and methods for fabrication of anagent, such as a therapeutic agent or an imaging agent, entrapped orencapsulated in a vehicle, capsule, particle, vector, or carrier inorder to maximize treatment and/or imaging of malignancy whileminimizing the adverse effects of treatment and/or imaging.Particularly, the systems and methods of the invention may form at leasta two-liquid electrified jet, as described in FIG. 1, using a singlecapillary tube for the production of capsules having entrapped orencapsulated agents. More particularly, the invention provides systemsand methods for producing capsules and particles with at least oneencapsulsted and/or entrapped agents, such as therapeutic agent, imagingagents, and other constituents.

In one embodiment of the invention, the capsules may serve multiplepurposes. First, the capsules may protect the encapsulated agent frombiological attack and degradation before it reaches the target site.Second, the capsules may prevent surface interaction of the agent frominducing undesired signal pathways and introduce the generation of acascade of proteins or hormones within the target cells thereby limitingthe agents' effectiveness.

The capsule(s) having the entrapped or encapsulated agents may be usedfor the treatment or imaging of cancer in a patient. For example, thecapsule may have an entrapped or encapsulated therapeutic agent for thetreatment of malignant cancers such as lymphoma, leukemia, sarcoma,mesothelioma, glioma, germinoma, and choriocarcinoma. Furthermore, thecapsule may have an entrapped or encapsulated imaging agent for thedetection of cancer. The capsule of the invention may also have acombination of entrapped or encapsulated therapeutic agents and imagingagents.

FIG. 2, which illustrates an embodiment of the invention, is a schematicshowing the general architecture for a capsule 200 with a distinct coreregion 202 and shell region 204. The core 202 may contain one or moreagents, such as therapeutic agents of biological or abiological origin.The shell may be made from one or more biocompatible polymers and maycontain entrapped domains 206 of one or more materials, which may beresponsible for the opening of the shell when the vesicle issubstantially in close proximity to or inside the target site, such asmalignant cells. Additionally, the capsule 200 may also serve as acontrast agent for imaging purposes. A biological or functional groups208 may be chemically or physically grafted onto the surface of theshell 204, which may facilitate transport by receptor or surface chargemediated endocytosis to and/or into the target site, such as malignantcells. Specifically, for example, malignant cells may over-expresscertain receptors and surface modification of the capsule 200 withfunctional groups 208 that selectively bind to the over-expressedreceptors on the surface of the malignant cells may improve transport ofthe capsule 200 to and/or into the malignant cells.

There is no requirement that capsule 200 is spherical, otherconfigurations and shapes, such as oblong, tubular, and ellipsoid mayalso be suitable for use in the invention. Additionally, capsule 200 maybe configured as an independent particle or may be configured to be aseries of capsules attached to each other in a chain-like fashion. Inparticular, if capsule 200 is configured in a chain-like series, thecapsules may have a length in the range of about 0.03 μm to about 30 μm.Moreover, capsule 200 may be also be effective if it has an oblongshape, tubular shape or ellipsoid shape, if the length such as a minoraxis in the case of an ellipsoid, or a diameter in the case of a tube,is sufficiently small to allow passage through the target site, such asa malignant cell membrane. The diameter or minor axis of the capsule maybe in a range of about 10 nm to about 1 μm.

According to one embodiment of the invention, the architecture of thecapsule may be based upon the mass ratio of shell material to corematerial and the composition and mass of the agents such as therapeuticagents, imaging agents, and other constituents, for example.Furthermore, the distinct core and shell regions may not be repaired.One or more agents, such as therapeutic or imaging agents may beentrapped and/or dispersed inside particles having at least onebiocompatible substance such as s biopolymer. The capsule may includeother additives, such as, a chemically functionalized surface to exploitreceptor biochemistry properties of malignant cells, dispersed particlesthat may aid in the breaking or segregation of the therapeutic and/or animaging agent carrying capsules once the capsules are substantiallyproximate to or inside the malignant cells.

According to an embodiment of the invention, the capsule may have ashell thickness in the range of about 10 nm to about 10 μm, and moreparticularly, in a range of about 20 nm to about 150 nm. In particular,the capsule shell has a thickness of about 20 nm. Moreover, the capsulemay have a size in the range of about 25 nm to about 25 μm, and moreparticularly, in a range of about 50 nm to about 500 nm. Specifically,the capsule has a size of about 250 nm.

In another embodiment, the capsule including at least one agent, such asa therapeutic agent or an imaging agent, may be transported into thetarget cells by processes such as endocytosis, transformation, ortransduction. Once the capsule enters the target cell, the action of thetherapeutic agent for treatment may be induced by chemical stress orexternal radiation. Once activated, the therapeutic agent may induce thedesired effect, such as apoptosis of the target cells. The therapeuticagents may be genetic material such as nucleic acid, RNA, DNA, bacteria,viruses, proteins, nucleic acid fragments, nucleic acid encoding a geneproduct, and abiological agents. Examples of the imaging agents mayinclude magnetic particles, photonic sensitive materials, radioactiveisotopes, and contrasting agents.

The capsule of the invention may have magnetic nanoparticles of lowtoxicity dispersed in it for external magnetic guidance of the capsule.In particular, the magnetic nanoparticles may include ferromagneticmaterials, such as metallic iron and certain metal oxides such atransition metal oxide, and rare earth based magnetic materials.Specifically, the magnetic nanoparticle may be a magnetite, such asFe₃O₄ having a size in the range of about 1 nm to about 300 nm, andspecifically, a size of about 10 nm. In addition to magneticnanoparticles, other materials may be dispersed within the capsule suchas materials sensitive to light or electric fields such as metallicsilver or gold, which may be used for imaging effects. Moreover, themetallic silver or gold may be used in combination with radioactiveemitters such as beta, gamma, or alpha emitters for imaging effects.

Referring to FIG. 3, which illustrates an embodiment of the invention,an upward flow EHD system 300 is shown for forming at least a two-fluidelectrified jet (FIG. 1) by using a single capillary tube. Here, system300 may include syringe pump 302 containing a core fluid, syringe pump304 containing a shell fluid, a fluid bath 306 containing shell fluid308, tube 310, collection plate 312, voltage source 314, electrode 316,core fluid supply tube 318, shell fluid supply tube 320, open end 322 oftube 310, fluid flow of shell fluid 324, collection zone 326, and plume328. As depicted in FIG. 3, syringe pump 302 containing the core fluidsupplies the core fluid via supply tube 318 to tube 310 which deliversthe core fluid upwards from its open end 322, while the shell fluid 308is delivered via wetting and capillary forces from the fluid bath 306surrounding the tube 310. The fluid flow of the shell fluid isdesignated as numeral 324. The electrode 316 may placed in severallocations, such as on tube 310, in fluid bath 306, the core fluid supplytube 318, or the shell fluid supply tube 320, for example. The electricpotential of the core fluid and the electric potential of the shellfluid may be the same.

The process throughput may be increased by making a device includingmore than one tube operating as tube 310, as described below. Dependingon the physical and chemical properties of the shell fluid 308 and thecore fluid delivered through tube 310, the height difference between theopen end 322 of tube 310 and the surface of shell fluid 308 in the fluidbath 306, the ratio of core and shell fluids in the upwards travelingelectrified liquid jet may be varied.

Moreover, in a further embodiment, a collector electrode 312 may beplaced above the surface of shell fluid 308 in fluid bath 306 in avariety of orientations. For example, if a flat metal surface is used ascollector electrode 312, the surface of the collector electrode may beoriented parallel or non-parallel to the surface of shell fluid 308 influid bath 306, or a different type of collector electrode configurationmay be selected, such as a rotating drum.

The plume 328 may contain same-charge droplets that repel each other andthus, travel toward collector electrode 312 with an ever-increasingcross-sectional area. The electric potential of the core fluid and theelectric potential of the shell fluid may be the same, due to theirintimate electrical contact. The electrical potential and the voltageare related quantities. The electrical potential may depend upon theseparation between the point at which a cone-jet region is formed, suchas region 326, and electrode collection plate 312. The separation mayhave a distance in a range of about 1.5 inches to about 15 inches.Moreover, the applied voltage may be in a range of about 0.5 kV to about35 kV.

Turning to FIG. 4, which illustrates another embodiment of theinvention, shell fluid 408, tube 310, collection plate 312, shell fluidflow 424, and an extractor 414 are shown. The extractor 414, which is anintermediate electrode, may be incorporated to aid in the formation ofthe two-fluid electrified jet. The extractor 414 may be an orifice 416made on a conductive body 418 and may be placed at a short distance fromthe open end 422 of tube 310. The extractor body may be biased at anintermediate electrical potential between those of tube 310 and thecollection plate 312.

In an alternate embodiment, the direction that the electrified two-fluidejected need not be upwards. Similar wetting by capillary forces oftubes or conduits delivering the core fluid may be effected to producedownward flows of the two-liquid electrified fluid jets. Electrifiedfluid or solid structures may be made to undergo total or partialelectrical discharge at a region after the extractor. This configurationmay increase the time of travel of the fluid or solid structures fromthe region where the electrified meniscus or menisci are formed, to thecollection zone, because a smaller electrical charge means decreasedmobility. The totally or partially discharged fluid or solid structuresmay be transported to a collection zone via use of gas flows.

The concept of wetting the external wall of a tube or conduit used todeliver the core fluid for a two-liquid electrified fluid jet may adoptother configurations. Referring to FIG. 5, which illustrates anembodiment of the invention, a tube 510 used to deliver the core fluid,a second tube 532 used to deliver the shell fluid, and the shell fluidflow 524 is shown. Besides flow rates, electric field strength and othervariables considerations, the distance between the outer wall of tube510, the open end of tube 532, and the angle 536 may be adjusted toensure adequate wetting the outer wall of tube 510 for formation of adesired two-fluid electrified menisci and electrified two-fluid jet.Adding two or more tubes or conduits serving as tube 532 to deliver theshell fluid may be considered a natural extension of the systemillustrated in FIG. 5. Flow rates for both the core and the shell fluidsmay be forced by the action of gravity i.e., by placing their respectivesupply reservoirs at an elevated distance with respect to the regionwhere the electrified two-fluid meniscus is formed, by the action ofmechanical or digital pumps, or driven by magnetic fields if one or bothfluids are magnetic. Once angle 536 and the number of tubes or conduitsserving the purpose of tube 532 may be taken as trivial degrees offreedom to produce two-fluid electrified liquid jets, a number of othernatural extensions to FIG. 5 may be realized.

For example, FIG. 6, which illustrates another embodiment of theinvention, shows one such natural extension to the principles in FIG. 5.In FIG. 6A, eight tubes or conduits 632 may be used to deliver the shellfluid. Tubes 632 may be oriented at an angle of about 90 degrees, whenthe angle is defined in the same way as angle 536 in FIG. 5 is defined.Tube 610 delivers the core fluid, which is supplied by the core fluidinlet 644, and an electrified, two-fluid meniscus is formed when boththe shell and the core fluids are accelerated by the action of anelectric field gradient toward a collection zone 626, or collectorelectrode, as previously described. In FIG. 6A, a shell fluid inlet port640 may be used to feed the shell fluid first into an encasement 642that envelops tube 610, and then through tubes 632. The number of tubes632 to be used may be based on factors such as, but not limited to, thephysical and chemical properties of the shell and core fluids, and theoperating variables, such as, but not limited to, the flow rates of thecore and shell fluids, and the electric field strength and its spatialdistribution. The distance between the open ends of tubes 610 and theopen end of tube 632, and the cross sectional areas of tubes 610 andtube 632, as shown in FIG. 6B, may also be determined based on operatingvariables, and the physical and chemical properties of the involvedfluids. Tubes 632 need not be in contact with each other.

In another embodiment, the concept of wetting the tube or conduitdelivering the core fluid may be extended to other practicalconfigurations. FIG. 7A, which illustrates an embodiment of theinvention, shows a configuration, where the number of tubes 710 may beused to deliver the core liquid from their open ends. Tubes 710 may besandwiched between two walls 714. The shell fluid 716 may then bedelivered through the space between adjacent tubes 710 (FIG. 7B providesthe cross section of FIG. 7A). By the action of an applied electricfield, electrified, two-fluid menisci and jets may be created.

Moreover, gap 718 and gap 720 may be adjusted or designed based uponoperating variables such as, but not limited to, flow rates alike twofluids involved and the electric field strength and its spatialdistribution. In particular, the closer electrified menisci are packedi.e., the smaller gap 720 is made, the higher the electric fieldrequired for forming the menisci. Unless the gas environment surroundingthe electrified menisci is not air, corona discharges may occur, andthese may compromise process continuity. Thus, a person of skill in theart would determine gap 720 based on these principles, on knowledge ofthe physical and chemical properties of the core and shell fluids, andthe process variables. For example, if the physical and chemicalproperties of the core and shell fluids and the values of the operatingvariables require gap 720 to be long enough to cause dripping of theshell fluid in between adjacent tubes 710, flow deflectors or bafflesmay be incorporated between adjacent tubes 710 to convey the outer fluidtowards tubes 710. Gap 718 may be determined by the wetting phenomenaoccurring between the outer walls of tubes 710 and the shell fluid 716.Additionally, an extractor electrode orifice may be placed at a shortdistance away from the corresponding two-fluid electrified meniscus, asdescribed above, and may be used to facilitate formation of two-fluidelectrified structures.

Referring to FIG. 8A, which illustrates an embodiment of the invention,shows that alignment of tubes 810 delivering the core fluid may differfrom the alignment shown in FIGS. 7A-7B. As shown in FIGS. 8A-8B, tubes810 may be sandwiched between a cylindrical encasement 812 and a rod814, and shell fluid 816 may be delivered through the space betweenadjacent tubes 810. The number of tubes delivering the core fluid inFIGS. 7A-7B and FIGS. 8A-8B may be determined based on the desiredprocess throughput. Likewise, the inner diameters of the tubesdelivering the core fluid 810 may be in the range of about 0.05 mm toabout 2.0 mm. The diameter of tubes 810 may also be based on theproperties of the involved fluids and the chosen operating variablessuch as, but not limited to, the electric field strength and its spatialdistribution, and the flow rates. The cross sectional areas of the tubesdelivering the core liquid 810 need not be circular, as other shapessuch as, but not limited to, ellipsoidal and polygonal can produceelectrified two-fluid menisci and jets, and the open, cross sectionalarea of the tube need not be oriented perpendicular to the axis of thetube. The open end of the tube may adopt other shapes such as, but notlimited to, conical or beveled.

Pacing side-by-side a plurality of the fixture shown in FIGS. 7A-7B toincrease product throughput may be desired to take advantage of thegeneral principle of wetting the outer wall of a conduit or tubedelivering the core fluid with the shell-forming fluid. Likewise, theprinciples of operation of the device shown in FIGS. 8A-8B may be usedto sandwich more than one circular layer of tubes or conduits deliveringthe core fluid between cylindrical encasements. FIGS. 7A-7B and FIGS.8A-8B are thus not limiting, as the surfaces that may be used tosandwich a plurality of tubes delivering the core fluid need not becylindrical or flat, since other surfaces with regular or irregularshapes may be used by the person skilled in the art.

In one embodiment of the invention, the concept of wetting the outerwall of a tube or conduit delivering fluids by the action forced flowand electric fields can be extended to produce multi-fluid electrifiedmenisci and jets. Referring FIG. 9, which illustrates an embodiment ofthe invention, shows the use of an arrangement comprising two coaxialtubes 910 and 916 to deliver core fluid 914 and shell fluid 912, and athird tube or conduit 916 used to deliver a third fluid. There arenatural extensions of the principles of creating an enveloping steam ofa third fluid via wetting of the external wall of the outer tube orconduit of the two-tube coaxial arrangement. By way of example, morethan two tubes with cylindrical cross sections may be arranged in acoaxial manner, to deliver more than two fluids via forced flows andelectric fields. Thus, if N is the number of such coaxial cylindricaltubes, (N+1) fluids can be delivered by wetting the outer wall of theoutermost cylindrical tube of the coaxial arrangement of N tubes, and bythe action of forced flows and electric fields, to form an electrified(N+1)-fluids meniscus and jet or jets.

FIG. 10, which illustrates yet another embodiment of the invention,depicts wetting the external walls of tubes or conduits carrying thecore fluid with the shell fluid. Tube 1002 carries the core fluid. Tube1004 feeds the shell fluid into the porous body 1006. Porous body 1006delivers the shell fluid via wetting of the external wall of tube 1002.a volume of the porous body 1006 may be made to envelop tube 1002, andto contact tube 1002 at a fraction of its external wall 1008. The choiceof the porous material 1006 may be dictated by the physical and chemicalproperties of the shell fluid. As one example, a latex sponge mayconstitute a suitable material to handle shell fluids comprising waterand other dissolved or suspended ingredients. The type of porous body1006 and the pore size distribution of porous body 1006 may be based onthe desired flow rates of the core and shell fluids, and the physicaland chemicals properties of the shell fluid. Examples of porousmaterials may include silicone based polymers, polyamide basedcross-linked polymers, melamine-formaldehyde based polymers, liquidparaffin, polyvinyl alcohol polymers, and poly(l-lactide) acid.

Moreover, a fixture comprising an arrangement of a plurality of tubes orconduits 1002 may be employed. The cross-sectional areas of the tubes1002 delivering the core liquid need not be circular, as other shapessuch as, but not limited to, ellipsoidal and polygonal, may produceelectrified two-fluid menisci and jets, and the open, cross sectionalarea of the tube need not be oriented perpendicular to the axis of thetube. The open end of the tube may adopt other shapes such as, but notlimited to, conical or beveled. Tube 1002 may be substituted by amulti-tube arrangement consisting of more than one tube with cylindricalcross arranged in a coaxial manner, as described above, to deliver morethan two fluids via forced flows and electric fields. The concept ofplacing an extractor electrode orifice at a short distance away from thecorresponding electrified fluids meniscus, as described above, may beused expand the system illustrated in FIG. 10 and to facilitateformation of multi-fluid electrified structures.

Turning to FIG. 11, which illustrates another embodiment of theinvention, the delivery of the shell fluid via wetting phenomena isdepicted. A reservoir 1102 for the core fluid may rotate by means of anymechanical device (not shown) while partially immersed in a bath 1104 ofthe shell fluid. The shell fluid may wet the external wall 1108 of thereservoir having the core fluid by the action of capillary forces andother physical phenomena. In the zone where shell fluid wetting of theexternal wall of reservoir 1102 occurs, at least one of orifices 1106,capillaries protruding the external wall of reservoir 1102, or otherconduit shapes may be made on the external wall 1108 of reservoir 1102to deliver the core fluid through the shell fluid film when an electricfield is applied between reservoir 1102 and a collection zone orcollector electrode. The distance between adjacent orifices 1106,capillaries or conduit shapes, as well as the rotational speed ofreservoir 1102, type of reservoir 1102 material, size and shape,immersion depth of reservoir 1102 into bath 1104, the number and size oforifices 1106, capillaries or conduit shapes, may be based on thephysical and chemical properties of the core and shell fluids and theoperating variables such as, but not limited to, applied voltage betweenreservoir 1102 and a collection zone or collector electrode.

In one embodiment, once a two-fluid electrified liquid jet is formed bythe methodology of the invention, a variety of fluids may be processedinto different shapes such as, but not limited to, hollow fibers andcapsules. A family of chemical synthesis methods known as sol-gel may beused to produce such structures from a two-fluid electrified jet. Forexample, one way of making structures with defined core and shellregions of different composition, requires that the shell fluid undergototal or partial solvent evaporation during the time of travel fromejection of the two-fluid electrified from the two-fluid electrifiedmeniscus to the collector electrode or collection zone. The precursorfor the solid shell may remain in solution until a critical fraction ofsolvent evaporates to the surroundings. The core fluid, depending on howit is formulated by the skilled person, may or may not yield a solidphase on collection at the collection zone.

The methodology of the invention may be employed to produce structureswith defined core and shell regions from a wide variety of fluids withdifferent chemical compositions. Additional functions such as, but notlimited to, magnetic properties may be added to the core or the shellfluids, or both, by formulating the chemistry of the said fluids.

For example, the shells of capsules containing a core fluid withdissolved protein may be made magnetic by suspending magnetite particlesin the range of about 100 nm to about 1 nm in the shell fluid precursorprior to processing using the methods described herein. The core fluidand the shell fluid chemistry may be formulated in such a way thatneither the core fluid nor the shell fluid undergoes solidificationbefore entering the droplet-in-droplet regime depicted in FIG. 3. Oncein the droplet-in-droplet regime, a critical amount of solventevaporation in the shell fluid it achieved, and capsules with a solidshell and a fluid core are collected in the collector electrode.

A variety of proteins, DNA material, cells and other biologicallyoriginated matter may be encapsulated using the methodology of theinvention. In particular, enzymes are a class of proteins that may serveas biocatalysts, and may be encapsulated in fluid cores such as, but notlimited to, pH buffed aqueous solutions, and the shells may be formedfrom sol-gel precursors having suspended magnetite particles. Themagnetite mass fraction in the shell may permit magnetic transport ofthe capsules once suspended in fluid media. The capsules containing theencapsulated enzymes may be suspended in a fluid containing reactantsand products for a reaction catalyzed by the encapsulated enzyme. If theshell is designed with pores large enough to allow diffusion through itof reactants and products for the reaction catalyzed by the encapsulatedenzyme but not as large so as to permit the enzyme to diffuse out of thecapsule, it is possible to recover the encapsulated enzyme with the aidof magnetic forces after the desired extent of reaction has beenachieved.

For example, in one embodiment of the invention, silicon alkoxidesol-gel chemistry may produce pores in the capsules in the range ofabout 0.5 nm to about 2.0 nm when controlled, which may be sufficient toallow diffusion of many commercially important reactants and products,but not to allow, for example, about a 50 kDa to about a 60 kDatransaminase enzyme to escape out of the sol-gel derived shell. Anatural extension of this concept is to design core fluid formulationscontaining substances of biological origin other than proteins such as,but not limited to DNA, DNA fragments, genes, and cells.

According to one embodiment, a magnetite nano-particle formulationcompatible with the silicon alkoxides based shell chemistry may yieldmagnetite may be coated with a silicon alkoxide layer having a size inthe range of about 2 nm to about 4 nm. Such a silicon alkoxide layermakes it ideal for firmly anchoring the magnetite to the shell solidphase via cross-linking sol-gel processes. A specific mass fraction ofmagnetite phase in the shell is in the range of about 0.1 to about 0.8.FIG. 12 is a photograph of the electrified liquid meniscus formed underan electric field and forced flow of the shell formulation that hasmagnetite particles added.

In another embodiment of this invention, the capsule suitable for thetreatment and/or imaging of malignant cancers may be fabricated byemploying electrospray. In general, the electrospray methodology mayinclude delivering a liquid containing the encapsulate(s) through asingle nozzle, capillary, conduit, or orifice, applying an electricalpotential between the nozzle and a collection region (herein after“counter-electrode”), forming a DC electrospray for the liquidcontaining the encapsulate(s), accelerating the DC electrospray dropletsto produce vesicles. The vesicle may be formed during the time of flightof the DC electrospray from its delivery region to the counter-electrodethrough a reaction between a suitable shell-forming monomer with liquidcomponents in the electrospray. The shell-forming monomer and the liquidcomponents do not compromise the integrity and function of theencapsulate(s). The capsules may be collected on the counter-electrodeare collected for storage and use.

Referring to FIG. 13A, which illustrates an embodiment of the invention,a schematic of the electrospray methodology of the invention isprovided. The electrospray methodology is based on a combination ofchemical reactions between the electrospray and it surrounding gasphase. In FIG. 13A, the electrospray nozzle 1302 and cloud 1304 aredepicted. Furthermore, the inlet 1306 and outlet port 1308, thecounter-electrode 1310, the shell-forming reaction volume 1312, and thehigh-voltage DC source 1314 are also shown. The inlet 1206 may fill theshell-forming region or chamber with a gas containing the shell-formingmonomer. The electrospray method of the invention offers the simplicityand scalability of single-liquid electrospray, thereby avoiding the useof multiple coaxial capillary nozzles altogether. Since electrospraysmay be produced at low Reynolds numbers i.e., under conditions of lowshear, the electrospray method of the invention produces true core-shellcapsules without the low-yield and high-shear process penalties ofemulsion-polymerization methods. Furthermore, the methodology alsoallows production of capsules much smaller than AC single-liquidelectrospray.

In one embodiment, the electrospray may be delivered into a region inspace whose gaseous environment has an adequate concentration ofshell-forming monomer. As shown in FIG. 13A, the electrospray method maybe made continuous or semi-continuous, depending on whether or not thesupply of electrosprayed encapsulate containing liquid, the supply ofmonomer-containing gas, and the removal of encapsulated matter are allmade continuous. Since the shell-forming process is bound to stop aftera certain shell thickness is achieved because the encased liquid corecannot deliver the components that react with the reactive monomers viadiffusion toward the outer, growing shell indefinitely, the shellthickness of the vesicles may be controlled by controlling a number ofprocess variables, such as but not limited to, monomer concentration,concentration of reactive species in the electrospray, temperature,time-of-flight of the electrospray, voltage, electrospray mean dropletsize, and monomer and electrospray chemical properties, for example.

In another embodiment of the invention, the encapsulate-containingelectrospray may be passed through a solution containing theshell-forming monomer, instead of a gas containing the said monomer.

another embodiment, the electrospray method of the invention may be madediscontinuous, and the time of suspension of the electrically chargedspray within the cyanoacrylate containing gaseous volumes may beextended by applying a high voltage AC potential within the saidshell-forming reaction volume, as shown in FIG. 13B. Referring to FIG.13B, the AC electrodes 1316, and the AC voltage source 1318 are shown.The fluid or solution containing the encapsulate may be first made intoan electrospray cloud as depicted in FIG. 13A, but during some timepoint in the electrospray process, the DC high voltage and the fluidflow of encapsulate containing the fluid or solution switched off, whilethe AC voltage is turned on. Charged droplets may then be made to engagein oscillatory motion between the two AC electrodes 1316, and thefrequency of the AC field is set so as to prevent the charge dropletsstriking one of the AC electrodes 1316. Turning off the AC high voltagesource 1318 while turning the DC voltage back on after a desired amountof time has elapsed, permits collection of the vesicles in the DCcounter-electrode 1310. By extending the time the electrospray remainssuspended within the zone in which the shell of the vesicles is formed,the reaction may be controlled and the shell-forming time over a muchlarger range than with the setup shown in FIG. 13A may be achieved. Anatural extension of the vesicle production scheme shown in FIG. 13B maybe to replace the gaseous environment in the shell forming chamber withan insulating fluid or solution bath containing the shell formingmonomer or monomers.

In an alternative embodiment of the invention, a single fluidelectrified jet may be employed, in the form of a solution or anemulsion to produce capsules having at least one entrapped agent. Anemulsion, as used herein, generally refers to a single fluid that may beused to generate the particles instead of capsules with the EHD methodas described above, using the same concentration ranges of thetherapeutic components and other components, and the same averageparticle sizes as those of the capsules described below. Furthermore,magnetic nanoparticles may also be dispersed in the particles.

For example, an aqueous emulsified phase containing at least one agent,such as a therapeutic anchor imaging agent, and stabilizing salts andpolyelectrolytes, may be dispersed in organic solutions containing atleast one biopolymer. The resulting emulsion includes a biphasic mediumwhich does not form distinct lasers on electrified jet formation. Theaqueous phase may be formulated in an identical manner as that of thecore fluid in the case of the capsules described above, and the organicphase containing the biopolymer or biopolymers, which may also includemagnetic nanoparticles and/or photonic particles formulated as that ofthe shell fluid of the capsules, as described above. Alternatively, asingle fluid electrified jet formulation using certain co-solvents suchas, but not limited to, dimethyl sulfoxide, leads to a homogeneousliquid mixture or solution comprising both the biopolymeric matrixprecursor or precursors, and the active ingredients and receptors. Thesingle electrified liquid jet approach may lead to entrapped, instead ofencapsulated, active agents.

According to one embodiment, the shell-forming monomers may includecompounds in the cyanoacrylate family, such as alkyl-substitutedcyanoacrylates, for example. Moreover, the encapsulate-containingformulations may be compounds that readily react with the cyanoacrylatemonomer to yield a skin of shell during the time-of-flight of theelectrospray toward the counter-electrode, without chemically alteringthe intended function of the encapsulate. In particular, theencapsulates may contain water. The encapsulates may be of biologicalorigin, such as nucleic acids, DNA, DNA fragments thereof, RNA, RNAfragments thereof, proteins, lipids, carbohydrates, and combinationsthereof and/or any modification thereof. Additionally, the encapsulatesmay include abiological therapeutic drugs, as described below.

According to one embodiment of the invention, the biopolymers that maybe suitable for producing the shell of the EHD-derived capsules include,without limitation, PLA, PLGA, chitosan, alkyl metacrylates, and otherbioabsorbable polymers such as PCL, starch, and co-polymers resultingfrom any combination thereof. Moreover, PEG may also be incorporated,either as polymer chains or as co-polymeric fragments to one or more ofthe other biopolymers, or functionalized with groups such as thio,carboxylic acid and amine groups. Additionally, other constituents maybe incorporated in the shell fluid, such as nanosized magnetiteparticles having a size in the range of about 1 nm to about 300 nm andsurfactants, prior to electrospray to yield a magnetically sensitive MRImaterial. Furthermore, nanosized photonic sensitive silver, gold,palladium or a combination thereof, having a size in the range of about1 nm to about 50 nm may be added to the shell precursor fluid prior toelectrospray to yield a photonic sensitive material.

According to another embodiment, core fluids suitable for themethodology of the invention may include aqueous phase with buffer saltsand polyelectrolytes such as phosphate buffers to stabilize the solutionstructure of therapeutic agents of biological origin, one or moretherapeutic agents such as, but not limited to, tumor necrosis factoralpha (TNF-α) protein, or TNF-α, encoding cDNA for TNF-α, or Egr-TNF,one or more substances capable of inducing the action of Egr-TNF suchas, but not limited to, Temozolomide, or TMZ, and chemotherapy agentssuch as, but not limited to, acridinyl anisidide, allopurinol,altretamine, aminoglutethimide, androgen, arsenic trioxide,asparaginease, azacitidine, antiangiogenesis, bleomycin, bortezomib,busulfan, capecitabine, carboplatin, carmustine, cetuximab,chlorambucil, cisplatin, cladribine, cyclophasphamide, dacarbazine,dactinomycin, daunorubiein, diethylstilbestrol, docetaxel, doxorubicin,epirubicin, estramustine, etanidazole, etoposide, floxuridine,fludarabine, 5-fluorouracil, flrosol DA, ftorafur, fulvestrant,gefitinib, gemcitabine, gemtuzumab ozogamicin, hydroxyurea, idarubicin,ifosfamide, interferon alpha, interleukin-2, irinotecan, levamisole,fomustine, mechloroethamine, melphalan, menogaril, methotrexate,methyl-CCNU, mitomycycin, mitoxantrone, oxaliplatin, paclitaxel,pemetrexed, pentostalin, picamycin, procarbazine, raltitrexed,steptoxocin, florafur, temozolomide, teniposide, thalidomide,thioguanine, thiotepa, topotecan, tositumomab, trimetreate, valspodar,vinblastine, vincristine, vindesine, vinorelbine, combinations thereof,and functional equivalents thereof.

Moreover, the capsules or particles of the invention may containchemotherapy-enhancing drugs, such as but not limited to, xcytrin, andchemotherapy protective drugs, such as but not limited to, amifostine.Certain radioactive elements may be suitable for the treatment and/orimaging of cancer, and in one embodiment of this invention, theradioactive elements can be incorporated in the electrospray generatedparticles. For example, Actinium-227 isotopic parent of Radium-223 is aso-called alpha emitter employed to treat metastases in the skeletonderiving from breast and prostate cancers, and for radioimmunotherapy,Copper-67 a gamma and beta emitter utilized in cancer radioimmunotherapyand lymphoma and colon and breast cancer diagnostics, Iodine-131, may beused for breast cancer treatment, leukemia, and lymphoma, usingradioimmunotherapy, thyroid function and disease (e.g., cancer) studies,as well as non-malignant thyroid ailments (e.g., hyperthyroidism),Iodine-125, may be useful for implants for breast and prostate tumors,radiolabeling, prostate cancer brachytherapy, osteoporosis detection,tracer drugs, imaging dianostics, mapping of brain receptors, braincancer therapy, interstitial radiation treatment, determination plasmavolume and flomerular filtration rate, deep vein thrombosis detection,Rhenium-186, useful for treatment and diagnosis of breast, colon, livercancers and lymphoma via radioimmunotherapy, rheumatoid arthritistherapy, and bone cancer pain relief, and Yttirium-91, which is agamma-emitting trace for Yttrium-90, which in turn is utilized forbreast cancer radioimmunotherapy, and also lymphoma, kidney, colon,lung, prostate, ovarian, pancreatic, and liver cancersradioimmunotherapy.

In a further embodiment, radioactive isotopes may incorporated into thecapsules of the invention, and their most frequent uses in medicine andrelated fields are: Cadmium-109 for general cancer detection,Actinium-225 and Thorium-229, the latter is the parent of Actinium-225and grandparent of Bismuth-213 which are alpha emitters used in cancerradioimmunotherapy, Bismuth-212, and Thorium-228, the latter is theparent of Bismuth-212 which is an alpha emitter employed in cancerradioimmunotherapy, Cobalt-60, which is a radiation source for cancerradiotherapy, for food and medical supplies irradiation, Copper-64, apositron emitter employed for cancer therapy, Dysprosium-166, employedin cancer radioimmunotherapy, Erbium-169, useful for small-jointrheumatoid arthritis therapy, Europium-152 and Europium-154, useful asradiation sources for medical supplies and food irradation,Gadolinium-153, useful for osteoporosis evaluation, Gold-198, useful forprostate and brain cancers, and for implants and intracavity treatmentof ovarian cancer, Holmium-166, useful for myeloma therapy in targetedskeletal therapy, ablation of bone marrow, radioimmunotherapy of cancer,and rheumatoid arthritis therapy, Iridium-192, useful for spinal cordand brain tumor treatment, brachytherapy, and treatment of blocked bloodvessels, Luterium-177, useful for treatment of blocked blood vessels andcancer radioimmunotherapy, Molybdenum-99, which is parent ofTechnetium-99m, which in turn is used for liver, brain, lungs, and heartimaging, and deep vein thrombosis detection, Osmium-194, useful forcancer radioimmunotherapy, Palladium-103, used in prostate cancertherapy, Platinum-195m, used in metabolism and viodistribution ofcisplatin studies, Phosphorus-32, used for treatment of leukemia andpolycythernia rubra vera (blood cell disease), bone cancer treatment anddiagnosis, treatment of pancreatic, colon and liver cancer, treatment ofblocked blood vessels and intracavity therapy, and diagnosis of surfacetumors, Phosphorus-33, useful for leukemia therapy, treatment anddiagnosis of bone disease, treatment of blocked blood vessels, andradiolabeling, Rhenium-188 useful for radioimmunotherapy, treatment ofrheumatoid arthritis, bone cancer pain relief, and prostate cancertherapy, Rhodium-105, used for cancer radioimmunotherapy, Samarium-145,used for treatment of ocular cancer, Samarium-153, useful for cancerradioimmunotherapy and bone cancer pain relief, Scandium-47, useful forbone cancer pain relief and cancer radioimmunotherapy, Selenium-75,useful for detection of hyperactive parathyroid glands, as radiotracerin brain evaluations, adrenal cortex imaging via gamma scintigraphy,detection of steroid producing tumors, and pacreatic scans,Strontium-85, useful for brain scanning and bone cancer detection,Strontium-89 useful for treatment of multiple myelona and bone cancerpain relief, Thulium-170, used as energy source for medical deviceimplants, and as a gamma source in blood irradiators, Tin-117m, used inbone cancer pain relief and cancer immunotherapy, Tungsten-188, which isthe parent of Rhenium-188, which in turn is used for cancer treatmentand diagnosis, rheumatoid arthritis treatment, and treatment of blockedblood vessels and bone cancer pain relief, Xenon-127, used in pulmonaryfunction evaluation, brain disorder imaging, and brain blood flowanalysis, Ytterbium-175, used in cancer radioimmunotherapy, andYttrium-90, which is obtained from Yttrium-89, and is useful for livercancer treatment. Metallic or nonmetallic nanoparticles such as Ga and Bwhich are used in neutron activation treatment can be co-encapsulatedwith therapeutic agents. The capsules or particles of this invention mayoptionally contain chemotherapy-enhancing drugs, such as but not limitedto, seytrin, and chemotherapy protective drugs, such as but not limitedto, amifostine.

In another embodiment, the capsules of the invention may include atleast one surface biochemical that may attach chemically or physicallyto enhance the affinity of the capsules for glioma cells, which mayinclude epidermal growth factor, or EGF. The epidermal growth factor mayeither be incorporated onto the surface of the capsules post EHDprocessing, or during EHD processing as part of the shell fluidprecursor. In particular, content range of EGF in the capsules andparticles made by the methodologies of the invention may be in the rangeof about 0 to about 0.02 wt %. Moreover, other surface biochemicals andchemicals may include, intraleukin-13, intraleukin-4, peripheral typebenzodiazephine, vascular endothelial growth factor, platelet-derivedgrowth factor, fibroblast growth factors, urokinase-type plasminogenactivator, folic acid and folic acid derivatives, neurotrophin growthfactor, somatostatin, eBSA, eHSA, protamine, insulin, ApoE derivedpeptide, polysorbate-80, OX-26, transferring, glucose, mannose, RMP-7,thiamine, combinations thereof and functional equivalents thereof. Thesurface biochemicals may be of human, animal or recombinant origin andhave a content in the range of about 0 to about 1 wt % for thenon-estrogen based surface biochemicals.

In a further embodiment, the capsules may include at least one surfacebiochemical that may attach chemically or physically to enhance theaffinity of the capsule for lymphoma, myeloma and leukemia such asVEGR-2, tositumomab, a monoclonal antibody which binds to the CD20receptor, and other antibodies and monoclonal antibodies in particularthose designes for other clusters of differentiation, such as but notlimited to, CD5, CD7, CD13, CD19, CD22, CD33, CD52, and CD61, andantimyeloperoxidase any combinations thereof, and functional equivalentsthereof. These biochemicals and their derivatives may either beincorporated onto the surface of the capsules post EHD processing, orduring EHD processing as part of the shell fluid precursor. The contentrange of the surface receptors in the capsules is in the range of about0.0 to about 1.0 wt %. Additional biochemicals and chemicals that mayincorporate into the capsule may include, tyrosine kinase receptors,including fibroblast growth factor receptor (FGFR-1), EGF, TGF, VEGF-A,urokinase receptor, interleuking 4 receptor, retinoic acid receptor,heparin-binding EGF-like growth factor, or HB-EGF, amphiregulin,epiregulin, and neuregulins, human epidermal growth factor 2 (HER-2) andfamily members, erbB2 and erbB1 receptor family, intraleukin-13 andfamily derivatives, platelet-derived growth factor, urokinase-typeplasminon activator, folic acid and folic acid derivatives, neurotrphingrowth factor, somatostatin, combinations thereof, and functionalequivalents thereof. The surface biochemicals may of human, animal orrecombinant origin, and may be present in a content range of about 0.0to about 1.0 wt %.

In a yet further embodiment, the capsules may include at least onesurface biochemical that may attach chemically or physically to enhancethe affinity of the capsule for breast cancer is estrogen. Estrogen andits chemical derivatives may either be incorporated onto the surface ofthe capsules post EHD processing, or during EHD processing as part ofthe shell fluid precursor. The content range of estrogen or itsderivatives in the capsules and particles may be in the range of about0.02 to about 0.4 wt %. Other surface biochemicals and chemicals thatmay be incorporated into the surface of the capsules and particles mayinclude human epidermal growth factor 2 (HER-2) and family memebers,erbB2 and erbB1 receptor family, intraleukin-13 and family derivatives,vascular endothelial growth factor, platelet-derived growth factor,fibroblast growth factors, urokinase-type plasminon activator, folicacid and folic acid derivatives, neurotrophin growth factor,somatostatin, combinations thereof, and functional equivalents thereof.The surface biochemicals may be of human, animal or recombinant origin,and may be in a content range of about 0 to about 1 wt %.

In another embodiment, the capsules may include surface biochemical thatmay attach chemically or physically to enhance the affinity of thecapsule for pancreatic cancer cells may the tyrosine kinase receptors,including firbroblast growth factor (FGFR-1), EGF, TGF, VEGF-A,urokinase receptor, interleukin 4 receptor, retinoic acid receptor,heparin-binding EGF-growth gactor, or HB-EGF, ampnhiregulin, epiregulin,neuregulins, and functional equivalents thereof. These biochemicals andtheir derivatives may either be incorporated onto the surface of thecapsules post EHD processing, or during EHD processing as part of theshell fluid precursor. The content of these surface receptors in thecapsules or particles may be in the range of about 0.0 to about 1.0 wt%. Additional surface biochemicals and chemicals that may beincorporated into the surface of the capsules and particles may includehuman epidermal growth factor 2 (HER-2) and family members, erB2 anderB1 receptor family, intraleukin-13 and family derivatives,platelet-derived growth factor, skomatostatin, combinations thereof, andfunctional equivalents thereof. The surface biochemicals may be ofhuman, animal or recombinant origin, and may be in a content range ofabout 0 to about 1 wt %.

Moreover, other surface biochemicals and chemicals of the said capsulesand particles may be fused in signal sequences, mitochondrial, nuclear,actin, and tubulin, golgi, plasma membrane, peroxisome, and may be in acontent range of about 0.0 to about 1.0 wt %.

For example, according to one embodiment of the invention, theconcentration of Paclitaxel in the capsules or particles may be in therange of aabout 10 to about 3200 u. Specifically, a concentration ofTNF-α protein in the capsules of particles may be in the range of about100 μg/mL to about 1,000 μg/mL. More spefically, and concentration ofEgr-TNF in the capsules or particles may be in the range of about 10μg/mL to about 300 μg/mL. The therapeutic effects may also be achievedwith capsules containing either one or two compounds of thethree-component, Paclitaxel, TNF-α, and Egr-TNF group. Narrow ranges ofthe mass fractions of the therapeutic components and other capsulescomponents, the capsules average size, average shell thickness, andnarrow core to shell average mass ratios may be selected based on thelocation and size of the malignancy, and use magnetic nanoparticledoping to aid in the guiding of the said capsules to the malignancy maynot be desired. For example, direct tumor injection, may be used inplace of magnetically assisted delivery, and electromagnetic irradiationof the malignancy in contact with the particles or capsules by themethodologies of the invention may be used to trigger disruption of theparticle with a concomitant release of therapeutic agents, to initiatebiochemical processes such as, but not limited to, malignant cell DNAdamage, or a combination of the said electromagnetic irradiation inducedprocesses. DNA markers containing pCMV-Luc+plasmid containing thecytomegalovirus (CMV) promoter of pcDNA3 inserted upstream to thefirefly luciferase of the pGL2-basic vector plasmid and TK renillalucerferase may also be used as markers.

In an alternate embodiment, convection enhanced delivery, or CED, may beused in place of magnetically assisted delivery, and electromagneticirradiation of the malignancy in contact with the particles or capsulesmade by the methodoligies of the invention may be used to triggerdisruption of the capsule or particle with a concomitant release oftherapeutic agents, to initiate biochemical processes such as, but notlimited to, malignant cell therapeutic agents, to initiate biochemicalprocesses such, but not limited to, malignant cell DNA damage, or acombination of the said electromagnetic irradiation induced processes.

The agents suitable for encapsulation or entrapment in the capsules ofthe invention should not be construed to be limited to agents suitablefor cancer treatment or imaging of malignancy, as described above,However, alternative agents may be suitable for entrapment orencapsulation may include anti-inflammatory compounds, anti-allergics,glucocorticoids, anti- infective agents, antibiotics, antifungals,antivirals, mucolytics, antiseptics, vasoconstrictors, wound healingagents, local anaestetics, peptides, and proteins.

Examples of potentially useful anti-inflammatory compounds areglucocorticoids, and non-steroidal anti-inflammatory agents such abetamethasone, beclomethasone, budesonide, circlesonide, dexamethasone,desoxymethasone, fluconolone acetonide, flucortin butyl, dycrocontisone,hyrocontisone-17-butyrate, predicarbate, 6-methylprednisolone aceponate,memetrasone furoate, elastane-, prostaglandin-, leukotriene,bradykinin-antagonits, non-steroidal anti-inflammatory drugs (NSAIDs),such as ibuprofen, indometacin, including any pharmaceutically acceptedsalts, esters, isomers, steroisomers, diasteromers, epimers, solvates orother hydrates, prodrugs, derivatives, or any other chemical or physicalforms of active compounds comprising the respective active moieties.

Examples of potentially useful antiallergic agents include theafore-mentioned glucocorticoids, and nedoeromil, cetrizin, loratidin,montelukast, roflumilast, ziluton, omalizumab, heparins and heparinoidsand other antihistimains, Azelastine, Cetirizin, Desloratadin, Ebastin,Fexofenadin, Levocetirizi, Loratidin.

Examples of anti-infective agents, whose class or therapweutic categoryis herein understood as comprising compounds which are effective againstbacterial, fungal, and viral infections, i.e. encompassing the classesof antimicrobials, antibiotics, antifungals, antiseptics, andantivirals, are penicillins, incuding benzylpenicillins,(penicillin-G-sodium, clemizone penicillin, benzathine penicillin G),phenozypenicillins (penicillin V, propicillin) aminobenzylpenicillinss(ampicillin, bacampicillin), acylaminopenicillins (azlocillin,mezlocillin, piperacillin, apalcillin), carbopenicillins (carbenicillin,ticarcillin, temocillin), isoxazolyl penicillins (oxacillin,cloxacillin, dicloraxacillin, flucloaxacillin) and amiidine penicillins(mecillinam); cephalosporins, including cefazolins (cefazolin,cefazedone); cefuroximes (cerufoxim, cefamdole, cefotiam), cefoxitins(cefoxitin, cefotetan, lamamoxef, flomoxef), cefotaximes (cefotaxime,ceftriaxone, cefmenoxime), ceftazidimes (ceftazime, cefpirome,cefepime), cefalexins (cefalexin, cefaclor, cefadroxil, cefadine,loracarbef, cefproxil) and cefiximes (cefixime, cefpodoxim, proxetile,cefuroxime axetil, cefetamet pivoxil, cefotiam hexetil), loracarbef,cefepim, clavulanic acid/amoxicillin, Ceftobiprole; synergists,including beta-lactamase inhibitors, such as clavulanic acid, sulbactam,and tazobactam; carbapenems, including imipenem, cilastin, meropenem,doripenem, tebipenem, ertapenem, ritipenam, and biapenem; monobactams,including aztreonam; aminoglycosides, such as apramycin, gentamicin,amikacin, isepamicin, abrekacin, tobramycin, netilimicin, spectinomycin,streptomycin, capreomycin, neomycin, paromoycin, and kanamycin;macrolides, including erythromycin, clarythromycin, roxithromycin,axithromycin, dithromycin, josamycin, spiramycin and telithromycin;gyrase inhibitors or fluroquinolones, including ciprofloxacin,gatilfloxacin, norfloxacin, ofloxacin, levofloxacin, pertloxacin,lomefloxacin, fleroxacin, garenoxacin, clinafloxacin, sitafloxacin,prulifloxacin, olamufloxacin, caderofloxacin, gemifloxacin,balofloxacin, trovafloxacin, and moxifloxacin; tetracyclins, includingtetracyclin, oxytetracyclin, rolitetracyclin, minocyclin, doxycycline,tigecycline and aminocycline; glycopeptides, including vancomycin,teicoplanin, ristocetin, avoparcin, oritavancin, ramoplanin, and peptide4; polypeptides, including plectasin, dalvavancin, daptomycin,oritavancin, ramoplanin, dalbavancin, telavancin, bacitracin,tyrothricin, neomycin, kanamycin, mupirocin, paromomycin, polymyxin Band colistin; sulfonamides, including sulfadiazine, sulfamethoxazole,sulfalene, co-trimoxazole, co-trimetrol, co-trimoxazine, andco-tetraxazine; azoles, including clotrimazole, oxiconazole, miconazole,ketoconazole, itraconazole, fluconazole, metronidazole, tinidazole,bifonazol, ravuconazol, posaconazol, voriconazole, and ornidazole andother antifungals including flucytosin, griseofluvin, tonoftal,naftifin, terbinafin, amorolfin, ciclopiroxolamin, echinocandins, suchas micafungin, caspofungin, anidulafungin; nitrofurans, includingnitrofurantoin and nitrofuranzone; -polyenes, including amphotercin B,natamycin, nystatin, flucocytosine; other antibiotics, includingtithromycin, lincomycin, clindamycin, oxazolindiones (linzezolids),ranbezolid, streptogramine A+B, pristinamycin aA+B, Virginiamycin A+B,dalfopristin/qiunupristin (Synercid), chloramphenicol, ethambutol,pyazinamid, terizidon, dapson, prothionamid, fosfomycin, fucidinic acid,rifampicin, isoniazid, cycloserine, terizidone, ansamycin, lysostaphin,iclaprim, mirocin B17, clerocidin, filgrastim, and pentamidine;antivirals, including aciclovir, ganciclovir, birivudin, valaciclovir,zidovudine, didanosin, thiacytidin, stavudin, lamivudin, zalcitabin,ribavirin, nevirapirin, delaviridin, trifluridin, ritonavir, saquinavir,indinavir, foscarnet, amantadin, polophyllotoxin, vidarabine,tromantadine, and proteinase inhibitors; plant extracts or ingredients,such as plant extracts from chamomile, hamamelis, echinacea, calendula,papain, pelargonium, essential oils, myrtol, pinen, limonen, cincole,thymol, mentol, tee tree oil, alpha-hederin, bisabolol, lycopodin,vitapherole; wound healing compounds including dexpantenol, allantoin,vitamins, hyaluronic acid, alpha-antitrypsin, anorganic and organic zincsalts/compounds, interferones (alpha, beta, gamma), tumor necrosisfactors, cytokines, interleukins.

Examples of potentially useful mucolytics are DNase, P2Y2-agonists(denufosol), heparinoids, guaifenesin, acetylcysteine, carbocysteine,ambroxol, bromhexine, lecithins, myrtol, and recombinant surfactantproteins.

Examples of potentially useful local anaesthetic agents includebenzocaine, tetracaine, procaine, lidocaine and bupivacaine.

Examples of potentially useful antiallergic agents include theafore-mentioned glucocorticoids, nedocromil. Examples of potentiallyuseful peptides and proteins include antibodies against toxins producedby microorganisms, antimicrobial peptides such as cecropins, defensins,thionins, and cathelicidins.

Also, immunmodulators including methotrexate, azothioprine, cyclosporineA, tacrolimus, sirolimus, rapamcyn, mycofenolate, mofetil, cytotaticsand metastasis inhibitors, alkylants, such as nimustine, melphanlane,carmustine, lomustine, cyclophosphosphamide, ifosfamide, trofosfamide,chlorambucil, busulfane, treosulfane, prednimustine, thiotepa;antimetabolites, e.g., cytarabine, fluoruracil, methotrexate,mercaptopurine, tioguanine; alkaloids, such as vinblastine, vincristine,vindesine; antibiotics, such as alcarubicine, bleomycine, dactinomycine,daunorubicine, doxorubicine, epirubicine, idarubicine, mitomycine,plicamycine; complexes of secondary group elements (e.g. Ti, Zr, V, Nb,Ta, Mo, W, Pt) such as carboplatinum, cis-platinum and metallocenecompounds such as titanocendichloride; amsacrine, dacarbazine,estramustine, etoposide, beraprost, hydroxycarbamide, mitoxanthrone,procarbazine, temiposide; paclitazel, iressa, zactima,poly-ADP-ribose-polymerase (PRAP) enzyme inhibitors, banoxantrone,gemcitabine, pemetrexed, bevacizumab, ranibizumab may be suitable forentrapment or encapsulation in the capsules of the invention.

In a further embodiment other compounds may include proteinaseinhibitors, such as a-antil-trypsin; antioxidants, such as tocopherols,glutathion; pituitary hormones, hypothalamic hormones, regulatorypeptides and their inhibiting agents, corticotropine, tetracosactide,choriogonandotropine, urofolitropine, urogonadotropine, saomatotropine,metergoline, desmopressine, oxytocine, argipressine, ornipressine,leuproreline, triptoreline, gonadoreline, buscreline, nafareline,godelerine, somatostatine; parathyroide gland hormones, calciummetabolism regulators, dihydrotachysterole, calcitronine, clodronicacid, etidronic acid; thyroid gland therapeutics; sex hormones and theirinhibiting agents, anabolics, androgens, estrogens, gestagenes,antiestrogenes; anti-migraine drugs, such as proxibarbal, lisuride,methysergide, dihydroergotamine, ergotamine, clonidine, pizotifene;hypnotics, sedatives, benzodiazepines, barbituates, cyclopyrrolones,imidazopyridines, antiepileptics, zolpidem, barbiturates, phenytoin,primidone, mesuximide, ethosuximide, sultiam, carbamazepin, valproicacid, vigabatrine; antiparkinson drugs, such as levodopa, carbidopa,benserazide, selegiline, bromocriptine, amantadine, tiapride;antiemetics, such as thiethylperazine, bromopride, domperidone,granisetrone, ondasetrone, tropisetrone, pyridozine; analgesics, such asbuprenorphine, fentanyl, morphine, codeine, hydromorphone, methadone,fenpipramide, fentanyl, piritramide, pentazocine, buprenorphine,nalbuphine, tilidine; drugs for narcosis, such as N-methylatedbarbiturates, thiobarbiturates, ketamine, etomidate, propofol,benzodiazepines, droperidol, halperidol, alfentanyl, sulfentanyl;antirheumatism drugs including tumor necrosis factor-alfa, nonsteroidalantiinflammatory drugs; antidiabetic drugs, such as insulin,sulfonylurea derivatives, biguanids, flitizols, glucagon, diazocid;cytokines, such as interleukines, interferones, tumor necrosis factor(TNF), colony stimulating factors (GM-CSF, G-CSF, M-CSF); proteins, e.g.epoetine, and peptides, e.g. parathyrin, somatomedin C; heparine,heparinoids, urokinases, streptokinases, ATP-ase, prostacycline, sexualstimulants, or genetic material.

The description and examples given above are merely illustrative and arenot meant to be an exhaustive list of all possible embodiments,applications or modification of the invention. Thus, variousmodifications and variations of the described methods and systems of theinvention will be apparent to those skilled in art without departingfrom the scope and spirit of the invention. Although the invention hasbeen described in connection with specific embodiments, it should beunderstood that the invention as claimed should not be unduly limited tosuch specific embodiments. Indeed, various modifications of thedescribed modes for carrying out the invention which are obvious tothose skilled in the material sciences, polymer sciences, or relatedfields are intended to be within the scope of the appended claims.

The disclosures of all references and publications cited above areexpressly incorporated by reference in their entireties to the sameextent as if each were incorporated by reference individually.

EXAMPLES Specific Example 1

This example describes an encapsulation of a protein solution using amodification of the shell formulation that consisted of a solution ofbovine serum albumin (BSA) in the presence of salts such as phosphatesto stabilize the pH of the solution. Confocal fluorescence microscopy(CFM) and BSA with a fluorescent tag was used for visualizing thecapsule. About 2 mg fluorescent protein was dissolved in about 1 mL ofphophate-buffer saline (PBS), type buffer that stabilized the acid-baseproperties of this aqueous solution to a pH equal to about 7.4.

The final BSA concentration was adjusted to about 3 μM. Silica sol wasused as the shell fluid precursor following the sol aging procedures,with addition of tert-amyl alcohol to a 50:50 volume ratio.

In general, for solution aging purposes, an acidified tetraethylorthosilicate solution in ethanol may be aged at about 80° C. for about4 to about 6 hours. Tert-amyl alcohol was added to increase thehycrophobicity of the shell fluid, which further prevented anysignificant mixing between core and shell fluids.

The core and shell flow rates were adjusted to about 0.025 and about0.75 mL/h, respectively. Adequate voltages for capsule design were inthe range of about 11 to about 12 kV, and the distance between the twofluid electrified meniscus and the collection zone was in the range ofabout 4 to about 14 cm. The collection zone, or collector electrode, wasa flat metal surface kept at ground electrical potential, whereas thetwo-fluid electrified meniscus were kept at a positive electrical biasin the range of about 5 to about 18 kV. FIG. 14 shows photographsobtained with the aid of the CFM technique, in which confinement of BSAwas evident when the instrument was focused to yield a fluorescentsignal in the core fluid region of the capsules. Referring to Figure 14,Panel I shows the unfocused fluorescent signal and Panel II shows thefocused fluorescent signal.

Specific Example 2

The sol-gel methods and process variables used to encapsulate fluorscentBSA in Specific Example 1 were slightly modified to encapsulate atransaminase. This enzyme was used to catalyze the following reaction:

D,L glutamine+glyoxylic acid→L-glutamine (left unreacted)+α-ketoderivative (from D-glutamine)+glycine.

Each enantiomeric form has the ability to rotate polarized light, and atechnique known as polarimetry may be used to follow chemical reactionsinvolving enantiomers as a function of time. The reactants, on the lefthand side of the reaction shown above, are basically a mixture with nooptical activity, since the D,L prefix stands for about a 50:50 mixtureof the D and L enantiomers of glutamine. The products, on the right handside of the reaction shown above, become enriched in unreactedL-glutamine with a concomitant time-dependent optical rotation signalthat may be tracked by polarimetry, because this particular transaminaseonly catalyzes reactions involving D enantiomers. The reaction wasbuffered to a about pH of 7.5 with PBS.

Since the specific rotation of the glutamine isomers was low in purewater, an experimental scheme was devised to both stop the reactionprior to polarimetry quantification at different reaction times, and toincrease the sensitivity of the analytical technique. This was achievedby addition of about 1.0 mL of 5 M HCl to an aliquot taken from thereactor, which denatures the enzyme. The enzyme catalyst either settlesat the bottom of the vial, or was removed by centrifugation, andacidification increased the sensitivity of the polarimetry technique byabout ½ order of magnitude relative to that observed in nearly neutralsolutions.

The biocatalytic test was run using about 50 mM concentrations of thesubstrates, and enzyme concentrations of about 0.5 mg/mL. FIG. 15 showsan example of the activity of this transaminase in both its encapsulatedstate, and free in solution, once optical rotation was converted to anequivalent D-(or L-) glutamine concentration via previous determinationof calibration curves.

Specific Example 3 Particles with 3.1 LACZ as DNA Marker

The therapeutic solution was prepared by mixing 3.1 LACZ in about 10 mMBis-Tris propane buffer aqueous solution containing about 1 wt % ofisoporpanol and about 2 mM of CaCl₂. The final concentration of 3.1 LACZwas 700 μg/mL.

A biopolymer solution was prepared by mixing two functionalizedbiopolymers in chloroform: (a) COOH-poly(ethylene glycol)-b-polylactide,or PPEG-b-PLA, with a molecular weight of 2000-b-1940 Da using samenomenclature, respectively, and a Mw/Mn=1.2; and (b)Poly(caprolactone)-SH, or PCSH, with a molecular weight of 5,000 Da anda Mw/Mn=1.5. This solution was doped with a solution of magnetiteparticles with an average diameter of about 15 mm. The weight percentcontents of PEG-b-PLA, PCSH, and magnetite particles in this solutionwere about 0.071 wt % about 0.058 wt %, and about 0.004 wt %respectively.

The therapeutic and biopolymer solutions were mixed and dimethylsulfoxide, or DMSO, was added to form a homogenous solution. Particleswith an average diameter in the range of aabout 0.250 to about 1 μm weremade produced, but smaller capsules may be made by varying the processvariables. Specifically, the flow rate used was about 0.150 mL/h, andthe external voltage was about 7 kV.

Specific Example 4 Encapsulation of PDs Red 2 NUC

The compound in this example was formed by the two-jet system depictedin FIG. 5. The core fluid solution was first prepared by mixing PDs red2 NUC and about 10 mM Bis-Tris propane aqueous solution containing about1 wt % of isopropanol and about 2 mM of CaCl₂. The final concentrationof PDs red 2 NUC in the core fluid solution was about 22.5 μg/mL.

The shell fluid solution was prepared by mixing two functionalizedbiopolymers in chloroform: (a) COOH-poly(ethylene gycol) b-polyactide,or PEF-b-PLA, with a molecular weight of 2000-b-1940 Da using samenomenclature, respectively, and a Mw/Mn=1.5. The weight percent contentsof PEG-b-PLA and PCSH in the shell fluid solution were about 0.29 wt %and about 0.31 wt %, respectively.

Capsules with an average diameter in the range of about 0.250 μm toabout 1 μm were produced. Specifically, the core and shell fluid flowrates used were about 0.050 and about 0.300 mL/h, respectively, and theexternal voltage was about 7 kV.

Capsules with a PDs red 2 NUC loading in the range of about 0.01 toabout 25% by weight of the added polymer may be made by this method byadjusting the concentration of the core fluid solution.

Specific Example 5 Encapsulation of Green Fluorescent Protein

The core fluid solution was prepared by mixing Green FluorescentProtein, or GFP, and about 10 mM Bis-Tris propane aqueous solutioncontaining about 1 wt % of isopropanol and about 2 mM of CaCl₂. Thefinal concentration of GFP in the core fluid solution was about 30μg/mL.

The shell fluid solution was prepared by mixing two functionalizedbiopolymers in chloroform; (a) COOH-poly(ethylene glycol)-b-polylactide,or PEG-b-PLA, with a molecular weight of 3000-b-1940 Da using samenomenclature, respectively, and a Mw/Mn=1.2; and (b)Poly(caprolaetone)-SH, or PCSH were about 0.32 wt % and about 0.31 wt %,respectively.

Capsules with an average diameter in the range of about 0.250 μm toabout 1 μm were made produces, but smaller capsules may be made byadjusting the process variables. Specifically, the core and shell fluidflow rates used were about 0.050 ml/h, respectively, and the externalvoltage was about 8 KV.

Capsules with a GFP loading in the range of about 0.01 to about 20% byweight of the added polymer were made by this method of adjusting theconcentration of the core fluid solution.

Specific Example 6 Particles with Doxorubicin with Folic Acid FunctionalGroups

The therapeutic solution is prepared by dissolving Doxorubicinhydrochloride, or DOXO, in dichloromethane. The concentration of DOXO inthe therapeutic solution is about 1,000 μg/mL.

A bioplymer solution is prepared by mixing two functionalizedbiopolymers in chloroform: (a) Folate-functionalized poly(ethyleneglycol), or Folate-PEG, with a molecular weight of 5,000 Da, and (b)Poly(caprolactone-SH, or PCSH, with a molecular weight of 5,000 Da and aMw/Mn=1.4. The weight percent contents of Folate-PEG and PCSH are about0.30 wt % and about 0.30 wt %, respectively.

The therapeutic and biopolymer solutions are mixed to form a homogenoussolution. Particles with an average diameter in the range of about 0.250μm to about 1 μm are produced, but smaller capsules may be made byadjusting the process variables. Specifically, the flowrates used are inthe range of about 0.050 to about 0.300 ml/h. The external voltage isabout 7.5 kV.

Particles with a DOXO loading in the range of about 0.01 to about 25% byweight of the added polymer may be made by this method by adjusting theconcentration of the core fluid solution.

Specific Example 7 Encapsulation of Doxorubicin

The core fluid solution were prepared by mixing doxorubicinhydrochloride, or DOXO in about 10 mM Bis-Tris propane aqueous solutioncontaining about 1 wt % of isopropanol and about 2 mM of CaCl₂. Thefinal concentration of DOXO in the core fluid solution was about 1,000μg/mL.

The shell fluid solution was prepared by mixing two functionalizedbiopolymers in chloroform: (a) COOH-poly(ethylene glycol)-b-polyactide,or PEG-b-PLA, with a molecular weight of 2000-b-1940 Da using samenomenclature, respectively, and a Mw/Mn=12.2; and (b)Poly(caprolactone)-SH, or PCSH, with a molecular weight of 5,000 Da anda Mw/Mn=1.5. The weight percent contents of PEG-b-PLA and PCSH wereabout 0.3 wt % and about 0.3 wt %, respectively.

Capsules with an average diameter in the range of about 0.250 μm toabout 1 μm were produced, but smaller capsules may be made by adjustingthe process variables. Specifically, the core and shell fluid flow ratesused were 0.050 and 0.300 ml/h, respectively, and the external voltagewas 8.5 kV.

Capsules with a DOXO loading in the range of about 0.01 to about 23% byweight of the added polymer may be made by this method by adjusting theconcentration of the core fluid solution.

Specific Example 8 Encapsulation of Cobalt Nanoparticles

The core fluid solution was prepared by mixing an aqueous solution ofcobalt nanoparticles, or NP-Co and a 10 mM Bis-Tris propane aqueoussolution containing about 1 wt % of isopropanol and about 2 mM of CaCl₂.The final concentration of NP-Co in the core fluid solution was in therange of about 1,000 to about 500 μg/mL.

The shell fluid solution was prepared by mixing two functionalizedbiopolymers in chloroform: (a) COOH-poly(ethylene glycol)-b-polylactide,or PEG-b-PLA, with a molecular weight of 2000-b-1940 Da using samenomenclature, respectively, and a Mw/Mn=1.2; and (b)Poly(caprolactone)-SH, or PCSH, with a molecular weight of 5,000 Da anda Mw/Mn=1.5. The weight percent contents of PEG-b-PLA and PCSH wereabout 0.30 wt % to about 0.30 wt %, respectively.

Capsules with an average diameter in the range of about 0.250 μm toabout 1 μm were produced, but smaller capsules may be made by adjustingthe process variables. Specifically, the core and shell fluid flow ratesused were in the range of about 0.050 to about 0.150 ml/h respectively,and the external voltage was about 8 kV.

Capsules with a nanoparticle (Np)-Co loading in the range of about 0.01to about 11% by weight of the added polymer may be made by this methodby adjusting the concentration of the core fluid solution.

Specific Example 9 Particles with Doxorubicin with EGF Functional Groups

The therapeutic solution is prepared by dissolving Doxorubicinhydrochloride, or DOXO, in dichloromethane. The concentration of DOXO inthe therapeutic solution is about 1,000 μg/mL.

A biopolymer solution is prepared by mixing two functionalizedbiopolymers in chloroform: (a) EGF-functionalized polyethylene glycol).pr EGF-PEG, with a molecular weight of 5,000 Da, and (b)Poly(caprolactone)-SH, with a molecular weight of 5,000 Da and aMw/Mn=1.5. The weight percent contents of EGF-PEG and PCSH are about0.30 wt % and about 0.30 wt % respectively.

The therapeutic and biopolymer solutions are mixed to form a homogenoussolution. Particles with an average diameter in the range of about 0.250μm to about 1 μm are produced, but smaller capsules may be made byadjusting the process variables. Specifically, the flowrates used are inthe range of about 0.050 to about 0.300 ml/h range. The external voltageis aabout 7.5 kV.

Particles with a DOXO loading in the range of about 0.01 to about 25% byweight of the added polymer may be made by adjusting the concentrationof the core fluid solution.

Specific Example 10 Particles with Luciferase as DNA Marker

The therapeutic solution is prepared by mixing Luciferase in a 10 mMBis-Tris propane buffer aqueous solution containing about 1 wt % ofisopropanol and about 2 mM of CaCl₂. The final concentration ofLuciferase is about 334.5 μg/mL.

A biopolymer solution is prepared by mixing two functionalizedbiopolymers in chloroform: (a) COOH-poly(ethylene glycol)-b-polylactide,or PEG-b-PLA, with a molecular weight of 2000-b-1940 Da using samenomenclature, respectively, and a Mw/Mn=1.2; and (b)Poly(caprolactone)-SH, or PCSH, with a molecular weight of 5,000 Da anda Mw/Mn=1.5. This solution is doped with a solution of magnetiteparticles with an average diameter of about 15 nm. The weight percentcontents of PEG-b-PLA, PCSH, and magnetite particles in this solutionare about 0.071 wt %, about 0.058 wt %,, and about 0.004 wt %respectively.

The therapeutic and biopolymer solutions are mixed and dimethylsolfoxide, or DMSO, is added to form a homogenous solution. Particleswith an average diameter in the range of about 0.250 to about 1 μm areproduced, but smaller capsules may be made by varying the processvariables. Specifically, the flow rate used was about 0.159 mL/h, andthe external voltage was about 7 kV.

Particles with a Luciferase loading in the range of about 0.01 to about63% by weight of the added polymer may be made by adjusting theconcentration of the Luciferase-containing solution.

Specific Example 11 Particles with TK Renilla Lucerase as DNA Marker

The therapeutic solution was prepared by mixing TK Renilla Luciferase inabout 10 mM Bis-Tris propane buffer aqueous solution containing about 1wt % if isoporpanol and about 2 mM of CaCl₂. The final concentration ofTK Renilla Luciferase was about 334.5 μg/mL.

A biopolymer solution was prepared by mixing two functionalizedbiopolymers in chloroform: (a) COOH-poly(ethylene glycol)-b-polyactide,or PEG-b-PLA, with a molecular weight of 2000-b-1940 Da using samenomenclature, respectively, and a Mw/Mn=1.5. This solution was dopedwith a solution of magnetite particles with an average diameter of about15 nm. The weight percent contents of PEG-b-PLA, PCSH, and magnetiteparticles in this solution were about 0.071 wt %, about 0.058 wt %, andabout 0.004 wt % respectively.

The therapeutic and biopolymer solutions were mixed and DMSO was addedto form a homogenous solution. Particles with an average diameter in therange of about 0.250 to about 1 μm were produced, but smaller capsulesmay be made by varying the process variables. Specifically, the flowrate used was about 0.150 mL/h, and the external voltage was aabout 7kV.

Particles with a TK Renilla Luciferase loading the range of about 0.01to about 63% by weight of the added polymer may be made by adjusting theconcentration of the TK Renilla Luciferase-containing solution.

Specific Example 12 Encapsulation of Cobalt Nanoparticles in CapsulesContaining EGF and Mitochondria Localization Vector

The core fluid solution is prepared by mixing an aqueous solution ofcobalt nanoparticles, or NP-Co and a 10 mM Bis-Tris propane aqueoussolution containing about 1.0 wt % of isopropanol and about 2 mM ofCaCl₂. The final concentration of NP-Co in the core fluid solution is inthe range of about 1,000 to about 500 μg/mL.

The shell fluid solution is prepared by mixing three functionalizedbiopolymers in chloroform: (a) COOH-poly(ethylene glycol)-b-polylactide,or PEG-b-PLA, with a molecular weight of 2000-b-1940 Da using samenomenclature, respectively, and a Mw/Mn=1.2; (b) Poly(caprolactone)-SH,or PCSH, with a molecular weight of 5,000 Da and a Mw/Mn=1.5; and (c)EGF-functionalized poly(ethylene glycol), or EGF-PEG, with a molecularweight of 5,000 Da. The weight percent contents of PEG-b-PLA, PCSH andEGF-PEG are about 0.30 wt %, about 0.30 wt % and about 0.10 wt %,respectively.

A buffer solution containing a plasmid subcellular localization vectortargeted to the michondria is added. The concentration of the vector inthe shell fluid solution is in the range of about 0.0 to about 1.0 wt %.DMSO is added to form a homogenous solution.

Capsules with an average diameter in the range of about 0.250 μm toabout 1 μm are produced but smaller capsules may be made by adjustingthe process variables. Specifically, the core and shell fluid flow ratesused were about 0;050 and about 0.150 ml/h, respectively, and theexternal voltage was about 8 kV.

Capsules with a Np-Co loading in the range of about 0.01 to about 11% byweight of the added polymer may be made by adjusting the concentrationof the core fluid solution.

Specific Example 13 Encapsulation of Chlorambucil in Capsules ContainingCD19 Functional Groups on the Shell

The core fluid solution was prepared by dissolving Chlorambucil in about10 mM Bis-Tris propane aqueous solution containing about 1 wt % ofisopropanol, about 2 mM of CaCl₂ and DMSO. The final concentration ofChlorambucil in the core fluid solution was in the range of about 2,000to about 500 μg/mL.

The shell fluid solution was prepared by mixing two functionalizedbiopolymers in chloroform: (a) COOH-poly(ethylene glycol)-b-PLA, with amolecular weight of 2000-b-1940 Da using same nomenclature,respectively, and a Mw/Mn=1.2; and (b) Poly(caprolactone)-SH, or PCSH,with a molecular weight of 5,000 Da and a Mw/Mn=1.5. The weight percentcontents of PEG-b-PLA and PCSH were about 0.05 wt % and about 0.05 wt %,respectively. The shell fluid solution was mixed with a solutioncontaining CD19 dissolved in a mixture of dichloromethane andpolyethylene oxide (MW=400-1000).

Capsules with an average diameter in the range of about 0.250 μm toabout 1 μm were produced, but smaller capsules may be made by adjustingthe process variables. Specifically, the core and shell fluid flow ratesused were about 0.050 and about 0.150 ml/h, respectively, and theexternal voltage was about 8 kV.

Capsules with a Chlorambucil loading in the range of about 0.02 to about20.0% by weight of the added polymer may be made by adjusting theconcentration of the core fluid solution. Capsules with a CD19 loadingin the range of about 5 to about 100 μg per mg of the added polymer maybe produced by adjusting the concentration of the shell fluid solution.

Specific Example 14 Encapsulation of Chlorambucil in Capsules ContainingCD20 Functional Groups

The core fluid solution was prepared by dissolving Chlorambucil in aabout 10 mM Bis-Tris propane aqueous solution containing about 1.0 wt %of isopropanol, about 2.0 mM of CaCl₂ and DMSO. The final concentrationof Chlorambucil in the core fluid solution was in the range of about2,000 to about 500 μg/mL.

The shell fluid solution was prepared by mixing two functionalizedbiopolymers in chloroform (a) COOH-poly(ethylene glycol)-b-polylactide,or PEG-b-PLA, with a molecular weight of 2000-b-1940 Da using samenomenclature, respectively, and a Mw/Mn=1.2; and (b)Poly(caprolactone)-SH, or PCSH, with a molecular weight of 5,000 Da anda Mw/Mn=1.5. The weight percent contents of PEG-b-PLA and PCSH wereabout 0.05 wt % and about 0.05 wt % respectively. Thee shell fluidsolution was mixed with a solution containing CD20 dissolved in amixture of dichloromethane and polyethylene oxide (MW=400-1000).

Capsules with an average diameter in the range of about 0.250 μm toabout 1 μm were generated, but smaller capsules may be made by adjustingthe process variables. Specifically, the core and shell fluid flow ratesused were about 0.050 and about 0.150 ml/h. respectively, and theexternal voltage was about 8 kV.

Capsules with a Chlorambucil loading in the range of about 0.01 to about10.0 % by weight of the added polymer may be made by adjusting theconcentration of the core fluid solution. Capsules with a CD20 loadingin the range of about 5 to about 100 μg per mg of the added polymer maybe made by adjusting the concentration of the shell fluid solution.

Specific Example 15 Encapsulation of Chlorambucil and HydrochloroquineSulfate in Capsules Containing CD19 and CD20 Functional Groups

The core fluid solution was prepared by dissolving Chlorambucil andHydrochloroquine sulfate, or HCQ, in about 10 mM Bis-Tris propaneaqueous solution containing about 1.0 wt % of isopropanol, 2 mM of CaCl₂and DMSO. The final concentrations of Chlorambucil and HCQ in the corefluid solution were in the range of about 2,000 to about 500 μg/mL.

The shell fluid solution was prepared by mixing two functionalizedbiopolymers in chloroform: (a) (COOH-poly(ethyleneglycol)-b-polylactide, or PEG-b-PLA, with a molecular weight of2000-b-1940 Da using same nomenclature, respectively, and a Mw/Mn=1.2;and (b) Poly(caprolactone)-SH, or PCSH, with a molecular weight of 5,000Da and a Mw/Mn=1.5. The weight percent contents of PEG-b-PLA and PCSHare 0.05 wt % and 0.05 wt %, respectively. The shell fluid solution wasmixed with a solution containing CD19 and CD29 dissolved in a mixture ofdichloromethane and polyethylene oxide (MW=400-1000).

Capsules with an average diameter in the range of about 0.250 μm toabout 1 μm were produced, but smaller capsules may be made by adjustingthe process variables. Specifically, the core and shell fluid flow ratesused were about 0.050 and about 0.300 ml/h, respectively, the core andshell fluid flow rates used were about 0.050 and about 0.300 ml/h,respectively, and the external voltage was about 8 kV.

Capsules with a Chlorambucil and HCQ loading in the range of about 0.01to about 20.0% each by weight of the added polymer may be made byadjusting the concentration of the core fluid solution. Capsules with aCD19 and CD20 loading in the range of about 5 to about 100 μg each permg of the added polymer may be made by adjusting the concentration ofthe shell fluid solution.

Specific Example 16 Encapsulation of Chlorambucil in Capsules ContainingCD19 Functional Group and Golgi Complex Localization Vector

The core fluid solution is prepared by dissolving Chlorambucil in about10 mM Bis-Tris propane aqueous solution containing about 1 wt % ofisopropanol, and 2 mM of CaCl₂ and DMSO. The final concentration ofChlorambucil in the core fluid solution is in the range of about 2,000to about 500 μg/mL.

The shell fluid solution is prepared by mixing two functionalizedbiopolymers in chloroform: (a) COOH-poly(ethylene glycol)-b-polylactide,or PEG-b-PLA, with a molecular weight of 2000-b-1940 Da using the samenomenclature, respectively, and a Mw/Mn=1.2; and (b)Poly(caprolactone)-SH, or PCSH, with a molecular weight of 5,000 Da anda Mw/Mn=1.5. The weight percent contents of PEG-b-PLA and PCSH are about0.05 wt % and about 0.05 wt %, respectively. The shell fluid solution ismixed with a solution containing CD19 dissolved in a mixture ofdichloromethane and polyethylene oxide (MW=400-1000).

A buffer solution containing a plasmid subcellular localization vectortargeted to the Golgi complex is added. The concentration of the vectorin the shell fluid solution is in the range of about 0 to about 1 wt %.DMSO is added to form a homogenous solution.

Capsules with an average diameter in the range of about 0.250 μm toabout 1 μm are produced, but smaller capsules may be made by adjustingthe process variables. Specifically, the core and shell fluid flow ratesused are about 0.050 and about 0.150 ml/h, respectively, and theexternal voltage is about 8 kV.

Capsules with a Chlorambucil loading in the range of about 0.01 to about20.0% by weight of the added polymer may be produced by adjusting theconcentration of the core fluid solution. Capsules with a CD19 loadingin the range of aabout 5 to about 100 μg per mg of the added polymer maybe made by adjusting the concentration of the shell fluid solution.

Specific Example 17 Particles Containing Iodine-125 and EGF FunctionalGroups

The therapeutic solution is prepared by dissolving Iodine-125 indichloromethane. The concentration of Iodine-125 in the therapeuticsolution is about 1,000 μg/mL.

A biopolymer solution is prepared by mixing two functionalizedbiopolymers in chloroform: (a) EGF-functionalized poly(ethylene glycol),or EGF-PEG, with a molecular weight of 5,000 Da, and (b)Poly(caprolactone)-SH, or PCSH, with a molecular weight of 5,000 Da anda Mw/Mn=1.5. The weight percent contents of EGF-PEG and PCSH are about0.30 wt % and about 0.30 wt %, respectively.

The therapeutic and biopolymer solutions are mixed to form a homogenoussolution. Particles with an average diameter in the range of about 0.250μm to about 1 μm are produced, but smaller capsules may be made byadjusting the process variables. Specifically, the flowrates used are inthe range of about 0.050 to about 0.300 ml/h. The external voltage isabout 7.5 kV.

Particles with an Iodine-125 loading in the range of about 0.01 and 25%by weight of the added polymer may be made by adjusting theconcentration of the core fluid solution.

Specific Example 18 Encapsulation of Chlorambucil in Capsules ContainingGa/Fe Nanoparticles and Epidermal Growth Factor Receptor

The core fluid solution is prepared by dissolving Chlorambucil in a 10mM Bis-Tris propane aqueous solution containing about 1.0 wt % ofisopropanol, about 2.0 mM of CaCl₂ and DMSO. The final concentration ofChlorambucil in the core fluid solution is in the range of about 2,0000to about 500 μg/mL.

The shell fluid solution is prepared by mixing two functionalizedbiopolymers in chloroform: (a) Epidermal growth factor-functionalizedpoly(ethylene glycol), or EGF-PEG, with a molecular weight of 5,000 Da,and (b) Poly(caprolactone)-SH, or PCSH, with a molecular weight of 5,000Da and a Mw/Mn=1.5. This solution is doped with a solution of Fe₃O₄ andGa₂O₃ nanoparticles with an average diameter of about 15 nm. The weightpercent contents of EGF-PEG, PCSH, Fe₃O₄ and Ga₂O₃ in the shell fluidsolution are about 0.71 wt %, about 0.058 wt %, about 0.004 wt % andabout 0.004 wt %, respectively.

Particles with an average diameter in the range of about 0.250 μm toabout 1 μm are produced, but smaller capsules may be made by adjustingthe process variables. Specifically, the flow rates used are in therange of about 0.050 to about 0.300 ml/h. The external voltage is about7.5 kV.

Particles with a Chlorambucil loading in the range of about 0.01 toabout 25.0% by weight of the added polymer may be made by adjusting theconcentration of the core fluid solution.

Specific Example 19 Encapsulation of Chlorambucil in Capsules ContainingGa/B/Fe Nanoparticles and Epidermal Growth Factor Receptor

The core fluid solution is prepared by dissolving Chlorambucil in about10.0 mM Bis-Tris propane aqueous solution containing about 1.0 wt % ofisopropaniol, about 2.0 mM of CaCl₂ and DMSO. The final concentration ofChlorambucil in the core fluid solution is in the range of about 2,000to about 500 μg/mL.

The shell fluid solution is prepared by mixing two functionalizedbiopolymers in chloroform: (a) Epidermal growth factor-functionalizedpoly(ethylene glycol), or EGF-PEG, with a molecular weight of 5,000 Daand a Me/Mn=1.5. This solution is doped with a solution of Fe₃O₄Ga₂O₃and B₂O₃ nanoparticles with an average diameter of about 15 nm. Theweight percent contents of EGF-PEG, PCSH, Fe₃O₄, Ga₂O₃ and B₂O₃ in theshell fluid solution are about 0.071 wt %, about 0.058 wt %, about 0.004wt %, about 0.004 wt % and about 0.004 wt %, respectively.

Particles with an average diameter in the range of about 0.250 μm toabout 1 μm are produced, but smaller capsules may be made by adjustingthe process variables. Specifically, the flowrates used are in the rangeof about 0.050 to about 0.300 ml/h. The external voltage is aabout 7.5kV.

Particles with a Chlorambucil loading in the range of about 0.01 toabout 25% by weight of the added polymer may be made by adjusting theconcentration of the core fluid solution.

Specific Example 20 Particles with Paclitaxel with Folic Acid FunctionalGroups

The therapeutic solution is prepared by dissolving Paclitaxel indichloromethane. The concentration of Paclitaxel in the therapeuticsolution is about 1,000 μg/mL.

A biopolymer solution is prepared by mixing two functionalizedbiopolymers in chloroform: (a) Folate-functionalized poly(ethyleneglycol), or Folate-PEG, with a molecular weight of 5,000 Da, and (b)Poly(caprolactone)-SH, PCSH, with a molecular weight of 5,000 Da and aMw/Mn=1.5. The weight percent contents of Folate-PEG and PCSH are about0.30 wt % and about 0.30 wt %, respectively.

The therapeutic and biopolymer solutions are mixed to form a homogenoussolution. Particles with an average diameter in the range of about 0.250μm to about 1 μm are produced, but smaller capsules may be made byadjusting the process variables. Specifically, the flowrates used are inthe range of about 0.50 to about 0.300 ml/h. The external voltage isabout 7.5 kV.

Particles with a Paclitaxel loading in the range of about 0.01 to about25% by weight of the added polymer may be made by adjusting theconcentration of the core fluid solution.

Specific Example 21 Capsules with Paclitaxel with Folic Acid FunctionalGroups

The core fluid solution is prepared by mixing Paclitaxel in DMSO andabout 10 wt % of γ-cyclodextrine in about 10 mM Bis-Tris propane aqueoussolution containing about 1 wt % of isopropanol and about 2 mM of CaCl₂.The final concentration of Paclitaxel in the core fluid solution isabout 20 μg/mL.

The shell fluid solution is prepared by mixing two functionalizedbiopolymers in chloroform: (a) Folate functionalized poly(ethyleneglycol), or Folate-PEG, with a molecular weight of 5,000 Da; and (b)Poly(caprolactone)-SH, with a molecular weight of 5,000 Da and aMw/Mn=1.5. The weight percent contents of Folate-PEG and PCSH are about0.30 wt % each.

Capsules with an average diameter in the range of about 0.250 μm toabout 1 μm are generated but smaller capsules may be made by adjustingthe process variables. Specifically, the core and shell fluid flow ratesare about 0.050 and about 0.300 ml/h, respectively, and the externalvoltage is about 7.5 kV.

Capsules with a Paclitaxel loading in the range of about 0.01 to about0.2 % by weight of the added polymer are made by adjusting theconcentration of the core fluid solution.

Specific Example 22 Particles and Paclitaxel

The therapeutic solution is prepared by mixing Paclitaxel indichloromethane. The concentration of Paclitaxel in the therapeuticsolution is about 1,000 μg/mL.

A biopolymer solution is prepared by mixing two functionalizedbiopolymers in chloroform: (a) COOH-poly(ethylene glycol)-b-polylactide,or PEG-b-PLA, with a molecular weight of 2000-b-1940 Da using samenomenclature, respectively, and a Mw/Mn=1.2; and (b)Poly(caprolactone)-SH, or PCSH, with a molecular weight of 5,000 Da anda Mw/Mn=1.5. The weight percent contents of PEG-b-PLA and PCSH are about0.3 wt %, respectively.

The therapeutic and biopolymer solutions are mixed to form a homogenoussolution. Particles with an average diameter in the range of about 0.250μm to about 1 μm are produced, but smaller capsules may be made byadjusting the process variables. Specifically, the flow rate is in therange of about 0.050 to about 0.300 ml/h, and the external voltage isabout 7.5 kV.

Particles with a Paclitaxel loading in the range of about 0.01 to about25% by weight of the added polymer may be made by adjusting theconcentration of the core fluid solution.

Specific Example 23 Encaspsulation of Paclitaxel

The core fluid solution is prepared by mixing Paclitaxel in DMSO andabout 10 wt % of γ-cyclodextrine in about 10 mM Bis-Tris propane aqueoussolution containing about 1 wt % of isopropanol and about 2 mM of CaCl₂.The final concentration of Paclitaxel in the core fluid solution isabout 20 μg/mL.

The shell fluid solution is prepared by mixing two functionalizedbiopolymers in chloroform: (a) COOH-poly(ethylene glycol)-b-polylactide,or PEG-b-PLA, with a molecular weight of 2000-b-1940 Da using samenomenclature, respectively, and a Mw/Mn=1.2; and (b)Poly(caprolactone)-SH, or PCSH, with a molecular weight of 5,000 Da anda Mw/Mn=1.5. The weight percent contents of PEG-b-PLA and PCSH are about0.3 wt % each.

Capsules with an average diameter in the range of about 0.250 μm toabout 1 μm are produced, but smaller capsules may be made by adjustingthe process variables. Specifically, the core and shell fluid flow ratesare about 0.050 and about 0.300 ml/h, respectively, and the externalvoltage is about 7 kV.

Capsules with a Paclitaxel loading in the range of about 0.01 to about0.2% by weight of the added polymer may be made by adjusting theconcentration of the core fluid solution.

Specific Example 24 Encapsulation of Gold Nanoparticles

The core fluid solution is prepared by mixing an aqueous solution ofgold nanoparticles, or NP-Au and a 10 mM Bis-Tris propane aqueoussolution containing about 1 wt % of isopropanol and about 2 mM of CaCl₂.The final concentration of NP-Au in the core fluid solution is in therange of about 1,000 to about 500 μg/mL.

The shell fluid solution is prepared by mixing two functionalizedbiopolymers in chloroform: (a) COOH-poly(ethylene glucol)-b-polylactide,or PEG-b-PLA, with a molecular weight of 2000-b-1940 Da using samenomenclature, respectively, and a Mw/Mn=1.5. The weight percent contentsof PEG-b-PLA and PCSH are about 0.30 wt % and about 0.30 wt %,respectively.

Capsules with an average diameter in the range of about 0.250 μm toabout 1 μm are produced, but smaller capsules may be made by adjustingthe process variables. Specifically, the core and shell fluid flow ratesare about 0.050 and about 0.150 ml/h, respectively, and the externalvoltage is about 8 kV.

Capsules with a gold nanoparticle loading in the range of about 0.01 toabout 11% by weight of the added polymer may be made by adjusting theconcentration of the core fluid solution.

Specific Example 25 Encapsulation of Silver Nanoparticles

The core fluid solution is prepared by mixing an aqueous solution ofsilver nanoparticles, or NP-Ag and a 10 mM Bis-Tris propane aqueoussolution containing about 1 wt % of isopropanol and about 2 mM of CaCl₂.The final concentration of NP-Ag in the core fluid solution is in therange of about 1,000 to about 500 μg/mL.

The shell fluid solution is prepared by mixing two functionalizedbiopolymers in chloroform: (a) COOH-poly(ethylene glycol)-b-polylactide,or PEG-b-PLA, with a molecular weight of 2000-b-1940 Da using samenomenclature, respectively, and a Mw/Mn=1.5. The weight percent contentsof PEG-b-PLA and PCSH are about 0.30 wt % and aabout 0.30 wt %,respectively.

Capsules with an average diameter in the range of about 0.250 μm toabout 1 μm are produced, but smaller capsules may be made by adjustingthe process variables. Specifically, the core and shell fluid flow ratesare about 0.050 and about 0.150 ml/h, respectively, and the externalvoltage is about 8 kV.

Capsules with a silver nanoparticle loading in the range of about 0.01to about 11% by weight of the added polymer may be made by adjusting theconcentration of the core fluid solution.

Specific Example 26 Encapsulation of Palladium Nanoparticles

The core fluid solution is prepared by mixing an aqueous solution ofpalladium nanoparticles, or Np-Pd and about 10 mM Bis-Tris propaneaqueous solution containing about 1 wt % of isopropanol and about 2 mMof CaCl₂. The final concentration of NP-Pd in the core fluid solution isin the range of about 1,000 to about 500 μg/mL.

The shall fluid solution is prepared by mixing two functionalizedbiopolymers in chloroform: (a) COOH-poly(ethylene glycol)-b-polylactide,or PEG-b-PLA, with a molecular weight of 2000-b-1940 Da using samenomenclature, respectively, and a Mw/Mn=1.2; and (b)Poly(caprolactone)-SH, or PCSH, with a molecular weight of 5,000 Da anda Me/Mn=1.5. The weight percent contents of PEG-b-PLA and PCSH are about0.30 wt % and about 0.30 wt %, respectively.

Capsules with an average diameter in the range of about 0.250 μm toabout 1 μm are produced, but smaller capsules may be made by adjustingthe process variables. Specifically, the core and shell fluid flow ratesare about 0.050 and about o.150 ml/h, respectively, and the externalvoltage is about 8 kV.

Capsules with a palladium nanoparticles loading in the range of about0.01 to about 11% by weight of the added polymer may be made byadjusting the concentration of the core fluid solution.

Specific Example 27 Particles with Paclitaxel and Estradiol FunctionalGroups

The therapeutic solution is prepared by mixing Paclitaxel indichloromethane. The concentration of Paclitaxel in the therapeuticsolution is about 1,000 μg/mL.

A biopolymer solution is prepared by mixing two functionalizedbiopolymers in chloroform: (a) Estradiol functionalized poly(ethyleneglycol), or EST-peg, with a molecular weight of 5,000 Da; and (b)Poly(caprolactone)-SH, or PCSH, with a molecular weight of 5,000 Da anda Me/Mn=1.5. The weight percent contents of EST-PEG and PCSH are about0.30 wt % each.

The therapeutic and biopolymer solutions are mixed to for a homogenoussolution. Particles with an average diameter in the range of about 0.250μm to about 1 μm are produced, but smaller capsules may be made byadjusting the process variables. Specifically, the flow rate is in therange of about 0.050 to about 0.300 ml/h range, and the external voltageis about 7.5 kV.

Particles with a Paclitaxel loading in the range of about 0.01 to about25% by weight of the added polymer may be made by adjusting theconcentration of the core fluid solution.

Specific Example 28 Capsules with Paclitaxel and Estradiol FunctionalGroups

The core fluid solution is prepared by mixing Paclitaxel in DMSO andabout 10 wt % of γ-cyclodextrine in about 10 mM Bis-Tris propane aqueoussolution containing about 1 wt % of isopropanol and 2 mM of CaCl₂. Thefinal concentration of Paclitaxel in the core fluid solution is about 20μg/mL.

The shell fluid solution is prepared by mixing two functionalizedbiopolymers in chloroform: (a) Estradiol functionalized poly(ethyleneglycol), or EST-PEG, with a molecular weight of 5,000 Da; and (b)Poly(caprolactone)-SH, with a molecular weight of 5,000 Da and aMw/Mn=1.5. The weight percent contents of EST-PEG and PCSH are about0.30 wt % each.

Capsules with an average diameter in the range of about 0.250 μm toabout 1 μm are produced, but smaller capsules may be made by adjustingthe process variables. Specifically, the core and shell fluid flow ratesare about 0.050 and about 0.150 ml/h, respectively, and the externalvoltage is about 8 kV.

Specific Example 29 Particles with Paclitaxel and Epidermal GrowthFactor Groups

The therapeutic solution is prepared by dissolving Paclitaxel indichlomethane. The concentration of Paclitaxel in the therapeuticsolution is about 1,000 μg/mL.

A biopolymer solution is prepared by mixing three functionalizedbiopolymers in chloroform: (a) COOH-poly(ethylene glycol)-b-polylactide,or PEG-b-PLA, with a molecular weight of 2000-b-1940 Da using samenomenclature, respectively, and a Mw/Mn=1.2; (b) Poly(caprolactone)-SH,or PCSH, with a molecular weight of 5,000 Da and a Mw/Mn=1.5; and (c)Epidermal growth factor-functionalized poly(ethylene glycol), orEGF-PEG, with a molecular weight of 5,000 Da. The weight percentcontents of PEG-b-PLA, PCSH and EGF-PEG are about 0.30 wt %, about 0.30wt % and about 0.10 wt %, respectively.

The therapeutic and biopolymer solutions are mixed to form a homogenoussolution. Particles with an average diameter in the range of about 0.250μm to about 1 μm are produced, but smaller capsules may be made byadjusting the process variables. Specifically, the flow rates are in therange of about 0.050 to about 0.300 ml/h. The external voltage is about7.5 kV.

Particles with a Paclitaxel loading in the range of about 0.02 to about25% by weight of the added polymer may be made by adjusting theconcentration of the core fluid solution.

Specific Example 30 Capsules with Paclitaxel and Epidermal Growth FactorGroups

The core fluid solution is prepared by mixing Paclitaxel in DMSO andabout 10 wt % of γ-cyclodextrine in about 10 mM Bis-Tris propane aqueoussolution containing about 1 wt % of isopropanol and 2 mM of CaCl₂. Thefinal concentration of Paclitaxel in the core fluid solution is about 20μg/mL.

The shell fluid solution is prepared by mixing three functionalizedbiopolymers in chloroform: (a) COOH-poly(ethylene glycol)-b-polylactide,or PEG-b-PLA, with a molecular weight of 2000-b-1940Da using samenomenclature, respective, and a Mw/Mn=1.2; (b) Poly(caprolactone)-SH, orPCSH, with a molecular weight of 5,000 Da and a Mw/Mn=1.5; and (c)Epidermal growth factor-functionalized poly(ethylene glycol), orEGF-PEG, with a molecular weight of 5,000 Da. The weight percentcontents of PEG-b-PLA, PCSH and EGF-PEG are about 0.30 wt %, about 0.30wt % and about 0.10 wt %, respectively.

Capsules with an average diameter in the range of about 0.250 μm toabout 1 μm are produced, but smaller capsules may be made by adjustingthe process variables. Specifically, the core and shell fluid flow ratesare about 0.050 and about 0.300 ml/h, respectively, and the externalvoltage is about 7.5 kV.

Capsules with a Paclitaxel loading in the range of about 0.01 to about0.2 % by weight of the added polymer are made by adjusting theconcentration of the core fluid solution.

Specific Example 31 Encapsulation of pCMV-Luc Plasmid

The core fluid solution is prepared by mixing a buffer solutioncontaining pCMV-Luc plasmid containing the cytomegalovirus (CMV)promoter of pcDNA3 inserted upstream to the firefly luciferase of thepGL2-basic vector plasmid with a 10 mM Bis-Tris propane aqueous solutioncontaining about 1 wt % of isopropanol and about 2 mM of CaCl₂. Thefinal concentration of pCMV-Luc in the core fluid solution is in therange of about 1 to about 1000 μg/mL.

The shell fluid solution is prepared by mixing two functionalizedbiopolymers in chloroform: (a) COOH-poly(ethylene glycol)-b-polylactide,or PEG-b-PLA, with a molecular weight of 2000-b-1940 Da using samenomenclature, respectively, and a Mw/Mn=1.2; and (b)Poly(caprolactone)-SH, or PCSH, with a molecular weight of 5,000 Da anda Mw/Mn=1.5. The weight percent contents of PEG-b-PLA and PCSH are about0.30 wt % and about 0.30 wt %, respectively.

Capsules with an average diameter in the range of about 0.250 μm toabout 1 μm are produced, but smaller capsules may be made by adjustingthe process variables. Specifically, the core and shell fluid flow ratesare about 0.050 and about 0.150 ml/h, respectively, and the externalvoltage is about 8 kV.

Capsules with a pCMV-Luc loading is in the range of about 0.01 to about11% by weight of the added polymer may be made by adjusting theconcentration of the core fluid solution.

Specific Example 32 Encapsulation of 425 TNF-α DNA and Bovine SerumAlbumin Rhodamine Conjugate

The core fluid solution was prepared by mixing 42S TNF-α DNA, and bovineserum albumin rhodamine conjugate, or BSA-rhodamine, in about 10 mMBis-Tris propane aqueous solution containing about 1 wt % of isopropanoland about 2 mM of CaCl₂. The final cconcentration of 425 TNF-α DNA andBSA-rhodamine in the core fluid solution were about 12.2 μg/mL, andabout 200 μg/mL, respectively.

The shell fluid solution was prepared by mixing an EGF graded to afluorescein-labeled poly(ethylene glycol)-NHS biopolymer, or F-PEG-EGF,with a molecular weight of 3,400 Da. The amount of F-PEG-EGF in theshell fluid solution was about 0.317 wt %.

Capsules with an average diameter of about 0.41 μm were produced, butsmaller capsules may be made by adjusting the process variables.Specifically, the core and shell fluid flow rates used were about 0.05and about 0.40 ml/h, respectively, and the applied voltage was about 6.5kV.

Capsules with a 425 TNF-α DNA loading in the range of about 0.01 toabout 30% by weight of the added polymer may be made by adjusting theconcentration of the core fluid solution.

Specific Example 33 Encapsulation of Temozolomide

The core fluid solution was prepared by mixing Temozolomide or TMZ inabout 0.1 M acetate buffer solution. The final concentration of TMZ inthe core fluid solution is about 10 μm.

The shell fluid solution was prepared by mixing two functionalizedbiopolymers in chloroform: (a) COOH-poly(ethylene glycol)-b-PLA, with amolecular weight of 2000-b-1940 Da using same nomenclature,respectively, and a Mw/Mn=1.2; and (b) Poly(caprolactone)-SH, or PCSH,with a molecular weight of 5,000 Da and a Me/Mn=1.5. The contents ofPEG-b-PLA and PCSH in the shell fluid solution were about 0.080 wt %,about 0.086 wt %, respectively.

Capsules with an average diameter of about 1.3 μm were produced, butsmaller capsules may be made by adjusting the process variables.Specifically, the core and shell fluid flow rates were about 0.05 andabout 0.30 ml/h, respectively, and the applied voltage was about 8.0 kV.

Capsules with a temozolomide loading in the range of about 0.01 to about2.4% by weight of the added polymer may be made by adjusting theconcentration of the core fluid solution.

Specific Example 34 Encapsulation of TNF-α Protein and Bovine SerumAlbumin Fluorescein Conjugate

The core fluid solution was prepared by mixing TNF-a protein, and bovineserum albumin fluorescein conjugate, or BSA-fluor 594, in about 10 mMBis-Tris propane aqueous solution containing about 1 wt % of isopropanoland about 2 mM of CaCl₂. The final concentrations of TNF-α protein andBSA-fluor in the core fluid solution were about 345 μg/mL and about 200μg/mL, respectively.

The shell fluid solution was prepared by mixing two functionalizedbiopolymers in chloroform: (a) COOH-poly(ethylene glycol)-b-polylactide,or PEG-b-PLA, with a molecular weight of 2000-b-1940 Da using samenomenclature, respectively, and a Mw/Mn=1.2; and (b)Poly(caprolactone)-SH, or PCSH, with a molecular weight of 5,000 Da anda Mw/Mn=1.5. The shell fluid solution is doped with a solution ofmagnetite particles with an average diameter of about 15 nm. Thecontents of PEG-b-PLA, PCSH, and magnetite particles in the shell fluidsolution were about 0.080 wt %, about 0.086 wt %, and about 0.006respectively.

Capsules with an average diameter of about 0.225 μm and about 0.550 μmwere produced, but smaller capsules may be made by adjusting the processvariables. Specifically, the core and shell fluid flow rates were about0.050 and about 0.300 ml/h, respectively, and the applied voltage wasabout 7.0 kV.

Capsules with a TNF-a protein loading in the range of about 0.01 toabout 23% by weight of the added polymer may be made by adjusting theconcentration of the core fluid solution.

The examples given above are merely illustrative and are not meant to bean exhaustive list of all possible embodiments, applications ormodifications of the invention. Thus, various modification and variationof the described methods and systems of the invention will be apparentto those skilled in the art without departing from the scope and spiritof the invention. Although the invention has been described inconnection with specific embodiments, it should be understood that theinvention as claimed should not be unduly limited to such specificembodiments. Indeed, various modifications of the described modes forcarrying out the invention which are obvious to those skilled in thepolymer sciences, molecular biology or related fields are intended to bewithin the scope of the appended claims.

The disclosures of all references and publications cited above areexpressly incorporated by reference in their entireties to the sameextent as if each were incorporated by reference individually.

1. An electrohydrodynamic system for producing a capsule having at leastone encapsulated agent, the system comprising: a hollow tube having anexternal wall and an interior configured to receive a core fluid, thehollow tube having an open discharge end configured to deliver the corefluid therethrough; a shell liquid source for delivering a shell liquidto the external wall of the hollow tube such that the open discharge endof the hollow tube is not surrounded by the shell liquid source; a corefluid supply tube arranged to supply the core fluid to the interior ofthe hollow tube; a shell liquid supply tube arranged to supply a shellliquid to the shell liquid source; a collector electrode positionedabove the shell liquid source configured to collect the producedcapsules; and an electric potential source to subject the core fluid andthe shell liquid to an electric potential to cause the core fluid andshell liquid to form a jet including an at least two-fluid electricallycharged fluid.
 2. The electrohydrodynamic system of claim 1 furthercomprising an extractor body positioned between the shell liquid sourceand the collector electrode.
 3. The system of claim 1, wherein the corefluid comprises at least one of a therapeutic agent and an imagingagent.
 4. The system of claim 1, wherein the hollow tube comprises aplurality of hollow tubes, the plurality of hollow tubes being locatedadjacent each other and forming a shape.
 5. An agent comprising thecapsule formed in the electrohydrodynamic system of claim
 1. 6. A methodfor fabricating a capsule having a core region and a shell region, themethod comprising: delivering a core fluid comprising at least one of atherapeutic agent and an imaging agent from a core fluid source throughan open discharge end of at least one hollow tube; delivering a shellliquid from a shell liquid source to an external wall of the at leastone hollow tube such that the open discharge ends of each of the atleast one hollow tubes is not surrounded by the shell liquid source;subjecting the core fluid and the shell fluid to an electric potentialremote from the open discharge ends of each of the at least one hollowtubes to cause the core fluid and shell liquid to form a jet includingan at least two-fluid electrically charged fluid; processing the jetinto a shape to form the capsule; and collecting the formed capsules ona collector.
 7. The method of claim 6, wherein the core fluid and theshell fluid is subjected to an applied voltage ranging from 0.5 kV to 35kV to form the jet including the at least two-fluid electrically chargedfluid.
 8. The method of claim 6 further comprising positioning anextractor between the open discharge ends of each of the at least onehollow tubes and the collector to aid in the formation of the at leasttwo-fluid electrically charged fluid.
 9. The method of claim 6, whereinsaid therapeutic agent is utilized for the treatment of malignantcancers.
 10. The method of claim 6, further comprising the step ofchemically or physically grafting a functional group on the surface ofthe shell region of the capsule.
 11. The method of claim 10, wherein thefunctional group is an entity selected from the group consisting ofhydroxyl groups, amino groups, carboxyl groups, carboxylic acidanhydride groups, mercapto groups, hydrosilicon groups, thio groups,carboxylic groups, amine groups, and any combination thereof.
 12. Themethod of claim 10, wherein the functional group is one or morecompounds selected from the group consisting of kinase receptors,fibroblast growth factor receptor, EGF, TGF, VEGF-A, urokinase receptor,interleukin-receptor, retinoic acid receptor, heparin-binding EGF-likegrowth factor, HB-EGF, amphiregulin, epireguin, neuregulins, andfunctional equivalents thereof.
 13. The method of claim 12, wherein thefunctional group has a content in the range of up to about 1 wt %. 14.The method of claim 10, wherein the functional group is capable ofchemically or physically attaching to glioma cells.
 15. The method ofclaim 10, wherein the functional group is epidermal growth factor and afunctional equivalent thereof.
 16. The method of claim 15, wherein thefunctional group has a content on the capsule in the range of up toabout 0.02 wt %.
 17. The method of claim 10, wherein the functionalgroup is capable of chemically or physically attaching to breast cancercells.
 18. The method of claim 10, wherein the functional group iscapable of chemically or physically attaching to at least one oflymphoma, myeloma and leukemia cancer cells.
 19. The method of claim 10,wherein the at least one hollow tube comprises a plurality of hollowtubes located adjacent to one another and forming a shape.
 20. A capsuleformed according to the method of claim
 10. 21-23. (canceled)